From 860687d8aedf31bdca55e70cc6a131239ab5fc40 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:07:33 +0100 Subject: [PATCH 01/36] First draft library prep RFC. --- rfcs/images/0000-lib_prep_rfc_fig1.png | Bin 0 -> 102822 bytes rfcs/images/0000-lib_prep_rfc_fig2.png | Bin 0 -> 108975 bytes rfcs/images/0000-lib_prep_rfc_fig3.png | Bin 0 -> 115425 bytes rfcs/images/0000-lib_prep_rfc_fig4.png | Bin 0 -> 180049 bytes rfcs/images/0000-lib_prep_rfc_fig5.png | Bin 0 -> 171584 bytes rfcs/images/0000-lib_prep_rfc_fig6.png | Bin 0 -> 140724 bytes rfcs/text/0000-rfc-library-preparation.md | 405 ++++++++++++++++++++++ 7 files changed, 405 insertions(+) create mode 100644 rfcs/images/0000-lib_prep_rfc_fig1.png create mode 100644 rfcs/images/0000-lib_prep_rfc_fig2.png create mode 100644 rfcs/images/0000-lib_prep_rfc_fig3.png create mode 100644 rfcs/images/0000-lib_prep_rfc_fig4.png create mode 100644 rfcs/images/0000-lib_prep_rfc_fig5.png create mode 100644 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z(ra^x`~VW{0DfD#;b^B&{Ew(a{;?5V{Gx4;v(xOA`fBfh zW5vvsC6ZS{YR%^7bp6is)zWsk^Q$Oyq{RNVO-n&bRg}1w-}C2Kw-G-oxa}BB@q6MF z`Cov1yU2M{dCM|p^7*WW{=83+s5e$vs;}>3CUz+~6Be5%tJ-!HV*m8>yfJX&%dmTh z|NF_f$4@K*tcPwRIo`WhDw_abaB9z;vF1JEg4g*&+*zEY7OQo5dM~c}0IT6Ij2)8(<_dr@k!G{}b~4%jOrqm&if2FaLM>N&F+g!KWdh$NNXL@YjLj zM+eXRznLNr9dTns^Iuyq5%H?}Q~Y4!N0^8QCv2Gh2=;%r=C4zdL333kt-pUJE&pP) zGzc*YTHELzrf9>8^jN9!m#^^GK>z>W{(YtSl~7KIZd-PR+LH^xuYf<|!ZJd+PhY(K EKg{J4N&o-= literal 0 HcmV?d00001 diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md new file mode 100644 index 00000000..fd67d7f4 --- /dev/null +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -0,0 +1,405 @@ +### DCP PR: + +***Leave this blank until the RFC is approved** then the **Author(s)** must create a link between the assigned RFC number and this pull request in the format:* + +`[dcp-community/rfc#](https://github.com/HumanCellAtlas/dcp-community/pull/)` + +# Representing sequencing library preparations in the HCA DCP metadata standard + +## Summary + +The current HCA DCP metadata model explicitly represents cell suspensions (single cells or multiple cells suspended in some media) but not the sequencing library preparations derived from them. This is creating challenges for contributors, consumers, and DCP implementation teams when submitting, processing, and interpreting sequencing data. + +Here we propose a solution for explicitly identifying library preparation biomaterials in a sequencing experiment by making them a first-class biomaterial type entity in the metadata schema alongside donor, cell suspension, imaged specimen, etc. + +## Author(s) + +*Recommended format for Authors:* + + `[Mallory Freeberg](mailto:mfreeberg@ebi.ac.uk)` + + `[Laura Clark](mailto:laura@ebi.ac.uk)` + + `[Name](mailto:username@example.com)` + +## Shepherd +***Leave this blank.** This role is assigned by DCP PM to guide the **Author(s)** through the RFC process.* + +*Recommended format for Shepherds:* + + `[Name](mailto:username@example.com)` + +## Motivation + +### Definitions + +**Logical unit** - a set of metadata and data files that are consumed together to provide a context for understanding, processing, and interpreting data. In the HCA DCP, a logical unit is instantiated by a primary or secondary “bundle”, although the meaning of bundle is not stable or clearly defined at the moment. + +**Library preparation** (biomaterial) - A collection of DNA fragments that have been prepared for sequencing from a single biological sample. DNA fragments can be prepared from genomic DNA or transcribed from RNA and are usually ligated to adapter and barcode oligonucleotides. + +**Library preparation protocol** - A protocol followed that converts endogenous RNA or DNA molecules into a form (cDNA) that can be sequenced. Often this protocol involves transcribing RNA to cDNA and ligating adapter and barcode oligonucleotides to the cDNA. + +### Problem background + +The current HCA DCP metadata standard explicitly represents cell suspensions: single or multiple cells suspended in some media after dissociation. In plate-based sequencing technologies, a single cell is isolated from other cells by being placed into a well on a plate, and then library preparation is carried out within each well. In droplet-based sequencing technologies, single cells are isolated using microfluidic partitioning (drops in oil) where barcoding and reverse transcription occur. Subsequently, the droplets are broken and cDNAs are pooled for amplification and library construction. + +From a cell suspension, one (Fig. 1A) or more (Fig. 1B) libraries can be prepared and sequenced. Each library preparation contains cDNA molecules representing a distinct and non-overlapping set of cells. The same library preparation can be sequenced more than once (Fig. 1C), each time producing a unique set of data files. For example, a library preparation can be sequenced on multiple flowcell lanes or a library preparation can be re-sequenced at a later time to generate more data. Regardless of how many times a library preparation was sequenced, **all of the sequence data files derived from one library preparation represent the same set of cells and therefore must be processed together**. Stated another way, **the logical unit of data should be based on a library preparation, all of the sequence files that come from it, and all of the biomaterials, protocols, and processes that generated it**. + + +![alt text](images/0000-lib_prep_rfc_fig1.png) + +**Figure 1**: Possible droplet-based sequencing experimental designs. A) An experiment where one library preparation was made from a cell suspension and then sequenced once. B) An experiment where two library preparations were made from the same cell suspension and then each library preparation was sequenced once. C) An experiment where two library preparations were made from the same cell suspension and then each library preparation was sequenced twice. Red boxes indicate the set of data files that need to be processed together. + +--- + +[Slides](https://drive.google.com/open?id=1vyw6N7qn24qBFAMoKL3nXLHcqpqYFq3Y) prepared by Nick Barkus (particularly slides 13-15) from the June 2019 DCP F2F describe why all sequence data files derived from one library preparation must be processed together. Briefly, a library preparation starts with a set of UMI barcodes attached to transcripts (1 barcode per transcript) and then everything gets amplified (potentially unevenly) and sequenced (Fig. 2A). Sequenced reads are potentially split between different sets of files if the library preparation is sequenced more than once. During processing, unique UMI barcodes are collapsed - meaning identical copies are only counted once - to reflect the original count (Fig. 2B). If files from the same library preparation are processed separately, a collapsed UMI barcode might appear in each set of files, thus inflating the count and leading to the wrong original count (Fig. 2C). + +![alt text](images/0000-lib_prep_rfc_fig2.png) + +**Figure 2**: Processing data files separately inflates UMI counts. A) Two transcripts - each with a unique UMI barcode - for gene A are amplified during library preparation, and the library preparation is sequenced twice to produce two sets of files. B) Files are processed together, and UMI barcodes are collapsed to produce a count of 2 for gene A. C) Files are processed separately, and UMI barcodes are collapsed separately to produce a count of 1 for gene A in the first set of files and 2 for gene A in the second set of files, resulting in an overall count of 3 for gene A (overestimation). + +--- + +### Current implementation + +The current HCA metadata standard does not represent a library preparation as an explicit biomaterial entity. At the moment, library preparations are represented by an *optional* metadata field ([`library_prep_id`](https://github.com/HumanCellAtlas/metadata-schema/blob/b8e48a3c7237010104a162c9013180172c2acaf5/json_schema/type/file/sequence_file.json#L81-L87)) on the sequence file entity that indicates which files come from the same library preparation. + +#### Data contributor perspective + +When data contributors supply metadata for their sequence files, they can enter a value for the `sequence_file.library_prep_id` field to indicate which sequence files were generated from the same library preparation. This value must be unique within the spreadsheet (and therefore project since 1 spreadsheet = 1 project), but it does not need to be globally unique (unique within the entire HCA DCP). + +| FILE NAME +(Required) | INPUT CELL SUSPENSION ID (Required) | LIBRARY PREPARATION ID (Optional) | +|:-|:-|:-| +| sequence_file.file_core.file_name | cell_suspension.biomaterial_core.biomaterial_id | sequence_file.library_prep_id| +| SRR7159837_1.fastq.gz | cell_suspension_1 | library_preparation_1 | +| SRR7159837_2.fastq.gz | cell_suspension_1 | library_preparation_1 | +| SRR7159838_1.fastq.gz | cell_suspension_1 | library_preparation_1 | +| SRR7159838_2.fastq.gz | cell_suspension_1 | library_preparation_1 | +| SRR7159839_1.fastq.gz | cell_suspension_1 | library_preparation_2 | +| SRR7159839_2.fastq.gz | cell_suspension_1 | library_preparation_2 | +| SRR7159840_1.fastq.gz | cell_suspension_1 | library_preparation_2 | +| SRR7159840_2.fastq.gz | cell_suspension_1 | library_preparation_2 | + +**Table 1**: An example set of sequence file metadata based on the experimental design in Fig. 1C. User-friendly field names in row 1. Fully-qualified programmatic field names in row 2. + +--- + +It is important to consider that, currently, if a valid spreadsheet with the metadata in Table 1 was submitted to the Ingestion Service and exported, it would create 1 bundle per row in Table 1. This is not correct because, at a minimum, _1/_2 file pairs must be in the same bundle to be processed together (and, e.g., R1/R2/I1 file triplets). A current workaround done by wranglers is to add a column in the Sequence file tab for the `process.process_core.process_id` field and enter the same value for the groups of files that should go in the same bundle. The solution proposed here for the library preparation problem also solves the problem of wranglers (and data contributors) having to fill in the `process_id` field in order to put the correct set of files in the same bundle. + +Assuming a data wrangler or data contributor has filled in the `process_id` field such that each process is assigned a unique identifier, anchoring a logical unit (bundle) on the ultimate process in the graph results in two bundles (Fig. 3A) which means that each bundle does not contain all the data files from the same library preparation. Ideally, logical units are anchored on the library preparation (Fig. 3B) such that all data files from the same library preparation are logically grouped together. + +![alt text](images/0000-lib_prep_rfc_fig3.png) + +**Figure 3**: Current and ideal grouping of logical units. A) Logical unit is anchored on ultimate process in the experimental graph, resulting in two bundles which each contain a subset of the data files produced from the same library preparation (not ideal). B) Logical unit is anchored on the library preparation entity, resulting in one logical unit that contains all the data files produced from the same library preparation (ideal). + +#### Data consumer perspective + +Converting the first row from Table 1 (and additional sequence file metadata for that row) to JSON produces: + +``` +{ + "describedBy": "https://schema.humancellatlas.org/type/file/9.0.0/sequence_file", + "schema_type": "file", + "file_core": { + "file_name": "SRR7159837_1.fastq.gz", + "format": "fastq.gz" + }, + "read_index": "read1", + "lane_index": 7, + "read_length": 26, + "library_prep_id": "library_preparation_1" +} +``` + +As described for Fig. 1C, all the sequence files derived from one library preparation must be processed together as a logical unit. In order to find all sequence files generated from the same library preparation, one must identify all the sequence files that (1) have the same value for `sequence_file.library_prep_id` AND (2) are from the same project (have the same project UUID) because `library_prep_id` is only unique within a project. This query is not ideal because (1) it is schema-aware, (2) it depends on an optional field, and (3) it queries over two variables (`library_prep_id` and project UUID) which makes the query slow. + +### Challenges + +The current implementation for representing sequence library preparations in the HCA DCP creates challenges for both HCA data contributors and consumers. Some of those challenges include: + +1. Data contributors are not required to indicate which data files represent sequencing the same library preparation more than once. Being able to do this is necessary to accurately represent the experimental design. +1. Data consumers find it challenging to determine which data files are from the same library preparation and therefore must be processed together. All data generated from one sequencing library preparation must be processed together. +1. Data consumers find it challenging to understand the relationship between cell suspensions from plate-based and droplet-based sequencing technologies because of granularity differences (1 cell versus many cells). This inconsistency creates confusion for data consumers when they are browsing or searching for data. + +### User Stories + +As a data contributor, I would like to be able to indicate which data files were generated from my library preparations so that I can represent my experiment accurately. + +As a data consumer, I would like to be able to identify which data files are from the same library preparation so that I can process all sequencing files which belong together at the same time. + +As a data consumer, I would like to be confident that data files from the same library preparation were processed together so that I know the expression matrices are accurate. + +## Scientific "guardrails" [optional] + +*Describe recommended or mandated review from HCA Science governance to ensure that the RFC addresses the needs of the scientific community.* + +TBD + +## Detailed Design + +### Overview + +We will resolve the challenges described above by **creating a first-class library preparation biomaterial entity** as part of the HCA metadata standard. This approach will allow the metadata model to explicitly represent which sequencing data files came from the same library preparation even if the data files were generated across multiple sequencing runs (i.e. multiple processes). This approach would also enable data consumers to more easily identify all data files associated with the same library preparation (i.e. from one logical unit) which is required for correct data processing. Unless and until the fundamental way that DNA sequencing experiments are performed changes, all sequencing-based experiments will involve generating library preparations; therefore, the addition of this entity to the metadata model will be used by all current and future sequence-based datasets. + +For cellular resolution experiments, the library preparation entity will have a cell suspension entity as input and sequence file entities as outputs. For bulk cell experiments, the library preparation entity will have a specimen (or organoid or cell line) entity as input and sequence file entities as outputs. Figure 3 below shows the current metadata model representing a droplet-based experiment sequencing a single cell suspension to produce two sets of two files (Fig. 4A). From this experimental design, it is not known whether the two sets of files represent one logical unit or two logical units. If the sequence files were produced from the same library preparation (Fig. 4B), then they represent one logical unit and must be processed together. If the sequence files were produced from different library preparation (Fig. 4C), then they represent two logical units and must be processed separately. + +![alt text](images/0000-lib_prep_rfc_fig4.png) + +**Figure 4**: Determining logical units from droplet-based experimental designs. A) An experiment modeled using the current metadata model which depicts four sequence files derived from one cell suspension. It is unclear what the logical units are. B) An experiment modeled using the proposed metadata model which depicts four sequence files derived from one library preparation. It is clear that these four files represent one logical unit (red outline). C) An experiment modeled using the proposed metadata model which depicts four sequence files derived from two library preparation. It is clear that these four files represent two logical units (one red outline, one blue outline). + +Arrows in graphs represent processes. Note that in B), two processes were used to derive the two sets of files. Both processes belong to this one logical unit. + +--- + +Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided. + +![alt text](images/0000-lib_prep_rfc_fig5.png) + +**Figure 5**: Determining logical units from plate-based experimental designs. A) An experiment modeled using the current metadata model which depicts six sequence files derived from three single cell suspension. Logical units are indicated (red, blue, and magenta outlines). B) An experiment modeled using the proposed metadata model which depicts six sequence files derived from three single cell library preparations. Logical units are indicated (red, blue, and magenta outlines). Arrows in graphs represent processes. + +**NB**: An experimental design where the same library preparation is sequenced more than once is not shown here. This is either (1) not even feasible or (2) highly unlikely to occur given that there isn’t a lot of material to sequence the same cell more than once. + +--- + +This proposal will require modifications to some DCP components (see “Implementation changes for DCP components” section below for more details). Importantly, with a new library preparation entity, the logical unit of data should be anchored on the library preparation, not the ultimate process used to generate a sequence file (colloquially referred to as the “assay process” in the Ingestion Service). + +### Metadata schema for library preparation entity + +**Standard biomaterial fields**. The new `library_preparation.json` schema will follow the schema structure indicated in the Metadata Schema Style Guide and will include the standard properties (fields) for biomaterial type schemas, including: + +``` +"describedBy": { + "description": "The URL reference to the schema.", +}, +"schema_version": { + "description": "The version number of the schema in major.minor.patch format.", +}, +"schema_type": { + "description": "The type of the metadata schema entity.", +}, +"provenance" : { + "description": "Provenance information provided by the system.", +}, +"biomaterial_core" : { + "description": "Core biomaterial-level information.", +} +``` + +Where the `biomaterial_core.json` schema contains the following properties: + +``` +"describedBy": { + "description": "The URL reference to the schema.", +}, +"biomaterial_id":{ + "description": "A [project-wide] unique ID for the biomaterial.", +}, +"biomaterial_name": { + "description": "A short, descriptive name for the biomaterial that need not be unique.", +}, +"biomaterial_description": { + "description": "A general description of the biomaterial.", +}, +"ncbi_taxon_id" : { + "description": "A taxonomy ID (taxonID) from NCBI.", +}, +"biosamples_accession": { + "description": "A BioSamples accession.", +}, +"insdc_sample_accession": { + "description": "An International Nucleotide Sequence Database Collaboration (INSDC) sample accession.", +}, +"supplementary_files": { + "description": "A list of filenames of biomaterial-level supplementary files.", +}, +"genotype": { + "description": "Genotype of the biomaterial.", +}, +"HDBR_accession": { + "description": "A Human Developmental Biology Resource (HDBR) sample accession.", +} +``` + +And the `provenance.json` schema contains the following properties (all supplied by Ingestion Service): + +``` +"describedBy": { + "description": "The URL reference to the schema.", +}, +"schema_version": { + "description": "The version number of the schema in major.minor.patch format.", +}, +"submission_date": { + "description": "When project was first submitted to database.", +}, +"submitter_id": { + "description": "ID of individual who first submitted project.", +}, +"update_date": { + "description": "When project was last updated.", +}, +"updater_id": { + "description": "ID of individual who last updated project.", +}, +"document_id": { + "description": "Identifier for document.", +}, +"accession": { + "description": "A unique accession for this entity, provided by the broker.", +} +``` + +The new `library_preparation.biomaterial_core.biomaterial_id` field will replace the function of the current `sequence_file.library_prep_id` field in the sequence file schema. Data contributors will be required to provide this field to ensure the correct experimental design is represented. It is unclear how the requirement will be enforced: at the moment there is no validation for the presence of a particular JSON type in a logical unit (i.e. bundle). + +The required fields for the library preparation entity will be: +- `library_preparation.biomaterial_core.biomaterial_id` +- `library_preparation.biomaterial_core.ncbi_taxon_id` + +**New library preparation-specific fields**. The new `library_preparation.json` schema will additionally include a field for capturing INSDC experiment accessions if the project is already archived. This field (`process.insdc_experiment.insdc_experiment_accession`) generally represents a single library preparation in the archives (ENA, SRA, DDJB), and currently lives awkwardly in a process module for historical reasons. Additional library preparation-specific fields could be added in the future. + +### Projection of library preparation entity in metadata spreadsheet + +Adding a new first-class biomaterial entity has the potential to add a lot of complexity to the metadata model. One area we want to avoid adding complexity is in the metadata spreadsheet, which is already quite complex for data contributors to fill out. Given that there are only two required (user-supplied) metadata fields proposed for library preparation entities, it is possible to keep the metadata spreadsheet relatively unchanged. + +Data contributors will supply a project-wide unique ID for each library preparation in the Sequence file tab using the `library_preparation.biomaterial_core.biomaterial_id` field in place of the current `sequence_file.library_prep_id` field. When the spreadsheet is imported, the Ingestion Service will generate a globally unique UUID for each library preparation entity. Unlike other biomaterials, there would not be a separate spreadsheet tab for library preparation entities for two reasons. 1. Adding a spreadsheet tab adds considerable complexity for data contributors. 2. There are only two fields that need to be supplied by the data contributor for which generating a whole new tab creates high overhead, especially considering one of the fields replaces a field already in the Sequence file tab. Without the ability to automatically infer `library_preparation.biomaterial_core.ncbi_taxon_id`, this field would need to be added to the Sequence file tab and would need to be filled in by contributors. For example: + + +| FILE NAME (Required) | INPUT CELL SUSPENSION ID (Required) | INPUT LIBRARY PREPARATION ID (Required) | NCBI TAXON ID (Required) | +|:-|:-|:-|:-| +| sequence_file.file_core.file_name | cell_suspension.biomaterial_core.biomaterial_id | library_preparation.biomaterial_core.biomaterial_id | library_preparation.biomaterial_core.ncbi_taxon_id | +| SRR7159837_1.fastq.gz | cell_suspension_1 | library_preparation_1 | 9606 | +| SRR7159837_2.fastq.gz | cell_suspension_1 | library_preparation_1 | 9606 | +| SRR7159838_1.fastq.gz | cell_suspension_1 | library_preparation_2 | 9606 | +| SRR7159838_2.fastq.gz | cell_suspension_1 | library_preparation_2 | 9606 | +| SRR7159839_1.fastq.gz | cell_suspension_1 | library_preparation_3 | 9606 | +| SRR7159839_2.fastq.gz | cell_suspension_1 | library_preparation_3 | 9606 | +| SRR7159840_1.fastq.gz | cell_suspension_1 | library_preparation_4 | 9606 | +| SRR7159840_2.fastq.gz | cell_suspension_1 | library_preparation_4 | 9606 | + +**Table 2**: An example set of sequence file metadata based on Fig. 1B experimental design. User-friendly field names in row 1; fully-qualified programmatic field names in row 2. Text in red indicates spreadsheet differences between Table 1 and Table 2. + +--- + +For the simplest experiments where a library preparation is sequenced only once (droplet- or plate-based), there will be a 1:1 relationship between `cell_suspension.biomaterial_core.biomaterial_id` and `library_preparation.biomaterial_core.biomaterial_id`. It might not be obvious to data contributors why they need to supply two different IDs; the importance will be clearly described in contributor-facing documentation for filling out the metadata spreadsheet. + +### Interpretation of library preparation entity by data consumers + +Data consumers will be able to find all data that need to be processed together by querying for all logical units (bundles) that are associated with a single library preparation entity as defined by its Ingestion Service-supplied UUID. For example, the query: + +``` +{ + "es_query": { + "query": { + "match": { + "files.library_preparation_0_json.provenance.document_id": "fd420633-6b3d-4361-ac0d-b57b8d9214d7" + } + } + } +} + +``` + +will return all bundles (primary and secondary) that contain the specified library preparation. + +Data consumers will not need to depend on multi-bundle notifications for processing all data files from the same library preparation.. All data files from a single library preparation will be put in the same logical unit if submitted at one time (in one submission envelope). Caveats: +1. Still unclear what the logical unit for an imaging experiments will be. More details under “Unresolved questions”. +1. Still unclear how to create logical units if data files from the same library preparation are submitted at different times (in more than one submission envelope). Is this a complex update to an existing logical unit? Or a new logical unit that re-uses many metadata files? + +Data consumers will benefit from the metadata model now aligning with INSDC “experiments” and “runs”. This alignment will also make archiving to ENA/SRA more straightforward. +- Library preparation is equivalent to an INSDC experiment +- Single set of files from a library preparation is equivalent to an INSDC run + + +| FILE NAME | INPUT LIBRARY PREPARATION ID | INSDC EXPERIMENT ACCESSION | INSDC RUN ACCESSIONS | +|:-|:-|:-|:-| +| sequence_file.file_core.file_name | library_preparation.biomaterial_core.biomaterial_id | process.insdc_experiment.insdc_experiment_accession | sequence_file.insdc_run_accessions | +| SRR7159837_1.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159837 | +| SRR7159837_2.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159837 | +| SRR7159838_1.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159838 | +| SRR7159838_2.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159838 | + +**Table 3**: An example set of sequence file metadata based on Fig. 1B experimental design showing how INSDC experiment accession aligns with library preparation ID and how INSDC run accession aligns with individual sets of files (i.e. a “run”). User-friendly field names in row 1; fully-qualified programmatic field names in row 2. **NB**: The `process.insdc_experiment.insdc_experiment_accession` field can be moved to the library preparation entity. + +--- + +### Implementation changes for DCP components + +#### Ingestion Service + +- Logical units will be anchored on a library preparation entity UUID (or possibly the process entity that is input to the library preparation entity) in the experimental graph (exporter). +- Schema template generator will be modified to (1) put the library preparation entity fields in the Sequence file tab, and (2) exclude all the library preparation entity fields except the required ones. +- Experimental graph (instantiated by links.json) will represent that (1) a library preparation protocol is followed to produce a library preparation from a cell suspension (or other biomaterial for bulk sequencing) and that (2) a sequencing protocol is followed to produce sequence file(s) from a library preparation. Users of the ingest-api will supply these links during programmatic submission. Otherwise, Ingestion Service (ingest-broker) will create the following links from metadata in the Sequence file tab and some graph assumptions (Fig. 6): + - Cell suspension <-> Process_1 <-> Library preparation <-> Process_0 <-> Sequence file + - Process_1 <-> Library preparation protocol + - Process_0 <-> Sequencing protocol + - Declare rules about linking + +![alt text](images/0000-lib_prep_rfc_fig6.png) + +Figure 5: Diagram of how protocol linking will change with new library preparation entity. A) Current metadata model showing that links to Sequencing protocol and Library preparation protocol entities are generated from the last process (arrow) in the graph. B) New metadata model showing that a link to the Sequencing protocol is generated from the ultimate process (process_0) in the graph, and a link to the Library preparation protocol is generated from the penultimate process (process_1) in the graph. + +#### Data Storage Service + +- No changes will be needed. +- Clients may need to update queries and subscriptions (some described below). + +#### Upload Service + +- No changes will be needed. + +#### Data Processing Pipelines + +- Subscription queries will reference a library preparation entity instead of cell suspension entity, where appropriate. +- If multiple-bundles:1-workflow remains a use case, pipelines can query for all “bundles” with a particular library preparation UUID in order to trigger a single workflow. + +#### Data Brower + +- Is anything coded against cell suspension? +- How will this affect displaying rows in “Sample” view? + +#### Matrix Service + +- Is anything coded against cell suspension? +- Will there be disruptions to how Matrix Service identifies single cells if we insert a library preparation entity between cell suspension and sequence file (thinking especially for plate-based technologies)? + +#### Query Service + +- Is anything coded against cell suspension? +- Will queries be made more difficult under this proposal? + + +### Acceptance Criteria [optional] + +- Data processing pipelines workflows can successfully be started and run on all data files from the same library preparation. +- Data consumers can find all data files associated with the same sequencing library preparation. +- Data contributors can complete a metadata spreadsheet without much more difficulty than before (how is this measured?). + +### Unresolved Questions + +#### To solve during RFC process +- How will the DCP enforce the requirement of having the library preparation entity in every logical unit (i.e. bundle)? +- How will current production sequencing datasets be updated? All of the current datasets will need to be updated as all of them will need to have library preparation entities added. This is a complex update - change in “bundle” structure by adding a library_preparation_0.json entity - so it can not be done with simple AUDR. If the update happens after GA but before complex AUDR, the UUIDs will have to be maintained somehow. + +#### To solve at future date + +- How might per-plate logical units be defined? The plate ID field is currently in the cell suspension schema. Creating a logical unit of “all data per plate” will not be made easier or harder if this proposal is accepted. +- Could this approach work in an analogous way for imaging experiments? I.e. anchor the logical unit on imaged specimen? It sounds like we don’t yet know yet what the “logical” unit is for image-based experiments. One option to anchor on the imaged specimen, but there are also other options. +- What are the naming conventions for user-supplied IDs and system-supplied, globally unique UUIDs? What are the rules for the names of the fields themselves? What are the rules for what values are valid for these fields? Related to metadata-schema issue [#733](https://github.com/HumanCellAtlas/metadata-schema/issues/733). + +### Drawbacks and Limitations [optional] + +Implementing the proposed design would require updating the current sequencing datasets in the production DSS to include a library preparation entity. This update would be a bundle structure update and would not be doable using the simple AUDR mechanism, but could possibly be done with complex AUDR. Updating primary bundles might affect downstream products (e.g. secondary bundles, matrices). + +This RFC is based on the current bundle structure and DCP design. If bundle structure or DCP design changes in the future (e.g. data processing pipelines can process more than one bundle with one workflow), the impact on library preparation modelling should be considered. Related to this [pre-RFC](https://docs.google.com/document/d/1akc8WrjxllNRkIAKcFD-8H3PsbrZQZjTZfgTI8KWuOI/edit#heading=h.s8wrvv1ovoyj). + +### Prior Art [optional] + +Previous [Library preparation entity RFC draft](https://docs.google.com/document/d/15-UkjbAlGDGhOPw1Zu09YdoaGw2Zeq5Y3uDlGsrz6pw/edit) +[Slides](https://drive.google.com/open?id=1vyw6N7qn24qBFAMoKL3nXLHcqpqYFq3Y) from June 2019 DCP F2F meeting outlining library preparation issue from data processing pipelines perspective. +[Slides](https://docs.google.com/presentation/d/1aV7Lrq8SWNxp3Jy98RxUh2SJrLiFPO4Y6AyaH3F1EUI/edit#slide=id.g599322258d_0_8) from June 2019 DCP F2F meeting outlining what is contained in bundles (logical units). + +### Alternatives [optional] + +The first attempt to clarify where library preparations fit in the experimental design was to add the optional `sequence_file.library_prep_id` field to the metadata standard. Having this field can “get the job done”, but at a high cost to consumers for needing to find it and difficulty for DCP components to use it to define “logical units” for co-processing. + From a6fab3b42a8207dd6bcac53daca49e1fcaf95244 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:09:17 +0100 Subject: [PATCH 02/36] Fixed image links. --- rfcs/text/0000-rfc-library-preparation.md | 12 ++++++------ 1 file changed, 6 insertions(+), 6 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index fd67d7f4..727079ff 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -46,7 +46,7 @@ The current HCA DCP metadata standard explicitly represents cell suspensions: si From a cell suspension, one (Fig. 1A) or more (Fig. 1B) libraries can be prepared and sequenced. Each library preparation contains cDNA molecules representing a distinct and non-overlapping set of cells. The same library preparation can be sequenced more than once (Fig. 1C), each time producing a unique set of data files. For example, a library preparation can be sequenced on multiple flowcell lanes or a library preparation can be re-sequenced at a later time to generate more data. Regardless of how many times a library preparation was sequenced, **all of the sequence data files derived from one library preparation represent the same set of cells and therefore must be processed together**. Stated another way, **the logical unit of data should be based on a library preparation, all of the sequence files that come from it, and all of the biomaterials, protocols, and processes that generated it**. -![alt text](images/0000-lib_prep_rfc_fig1.png) +![Figure 1](../images/0000-lib_prep_rfc_fig1.png) **Figure 1**: Possible droplet-based sequencing experimental designs. A) An experiment where one library preparation was made from a cell suspension and then sequenced once. B) An experiment where two library preparations were made from the same cell suspension and then each library preparation was sequenced once. C) An experiment where two library preparations were made from the same cell suspension and then each library preparation was sequenced twice. Red boxes indicate the set of data files that need to be processed together. @@ -54,7 +54,7 @@ From a cell suspension, one (Fig. 1A) or more (Fig. 1B) libraries can be prepare [Slides](https://drive.google.com/open?id=1vyw6N7qn24qBFAMoKL3nXLHcqpqYFq3Y) prepared by Nick Barkus (particularly slides 13-15) from the June 2019 DCP F2F describe why all sequence data files derived from one library preparation must be processed together. Briefly, a library preparation starts with a set of UMI barcodes attached to transcripts (1 barcode per transcript) and then everything gets amplified (potentially unevenly) and sequenced (Fig. 2A). Sequenced reads are potentially split between different sets of files if the library preparation is sequenced more than once. During processing, unique UMI barcodes are collapsed - meaning identical copies are only counted once - to reflect the original count (Fig. 2B). If files from the same library preparation are processed separately, a collapsed UMI barcode might appear in each set of files, thus inflating the count and leading to the wrong original count (Fig. 2C). -![alt text](images/0000-lib_prep_rfc_fig2.png) +![Figure 2](/rfcs/images/0000-lib_prep_rfc_fig2.png) **Figure 2**: Processing data files separately inflates UMI counts. A) Two transcripts - each with a unique UMI barcode - for gene A are amplified during library preparation, and the library preparation is sequenced twice to produce two sets of files. B) Files are processed together, and UMI barcodes are collapsed to produce a count of 2 for gene A. C) Files are processed separately, and UMI barcodes are collapsed separately to produce a count of 1 for gene A in the first set of files and 2 for gene A in the second set of files, resulting in an overall count of 3 for gene A (overestimation). @@ -89,7 +89,7 @@ It is important to consider that, currently, if a valid spreadsheet with the met Assuming a data wrangler or data contributor has filled in the `process_id` field such that each process is assigned a unique identifier, anchoring a logical unit (bundle) on the ultimate process in the graph results in two bundles (Fig. 3A) which means that each bundle does not contain all the data files from the same library preparation. Ideally, logical units are anchored on the library preparation (Fig. 3B) such that all data files from the same library preparation are logically grouped together. -![alt text](images/0000-lib_prep_rfc_fig3.png) +![Figure 3](images/0000-lib_prep_rfc_fig3.png) **Figure 3**: Current and ideal grouping of logical units. A) Logical unit is anchored on ultimate process in the experimental graph, resulting in two bundles which each contain a subset of the data files produced from the same library preparation (not ideal). B) Logical unit is anchored on the library preparation entity, resulting in one logical unit that contains all the data files produced from the same library preparation (ideal). @@ -144,7 +144,7 @@ We will resolve the challenges described above by **creating a first-class libra For cellular resolution experiments, the library preparation entity will have a cell suspension entity as input and sequence file entities as outputs. For bulk cell experiments, the library preparation entity will have a specimen (or organoid or cell line) entity as input and sequence file entities as outputs. Figure 3 below shows the current metadata model representing a droplet-based experiment sequencing a single cell suspension to produce two sets of two files (Fig. 4A). From this experimental design, it is not known whether the two sets of files represent one logical unit or two logical units. If the sequence files were produced from the same library preparation (Fig. 4B), then they represent one logical unit and must be processed together. If the sequence files were produced from different library preparation (Fig. 4C), then they represent two logical units and must be processed separately. -![alt text](images/0000-lib_prep_rfc_fig4.png) +![Figure 4](images/0000-lib_prep_rfc_fig4.png) **Figure 4**: Determining logical units from droplet-based experimental designs. A) An experiment modeled using the current metadata model which depicts four sequence files derived from one cell suspension. It is unclear what the logical units are. B) An experiment modeled using the proposed metadata model which depicts four sequence files derived from one library preparation. It is clear that these four files represent one logical unit (red outline). C) An experiment modeled using the proposed metadata model which depicts four sequence files derived from two library preparation. It is clear that these four files represent two logical units (one red outline, one blue outline). @@ -154,7 +154,7 @@ Arrows in graphs represent processes. Note that in B), two processes were used t Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided. -![alt text](images/0000-lib_prep_rfc_fig5.png) +![Figure 5](images/0000-lib_prep_rfc_fig5.png) **Figure 5**: Determining logical units from plate-based experimental designs. A) An experiment modeled using the current metadata model which depicts six sequence files derived from three single cell suspension. Logical units are indicated (red, blue, and magenta outlines). B) An experiment modeled using the proposed metadata model which depicts six sequence files derived from three single cell library preparations. Logical units are indicated (red, blue, and magenta outlines). Arrows in graphs represent processes. @@ -335,7 +335,7 @@ Data consumers will benefit from the metadata model now aligning with INSDC “e - Process_0 <-> Sequencing protocol - Declare rules about linking -![alt text](images/0000-lib_prep_rfc_fig6.png) +![Figure 6](images/0000-lib_prep_rfc_fig6.png) Figure 5: Diagram of how protocol linking will change with new library preparation entity. A) Current metadata model showing that links to Sequencing protocol and Library preparation protocol entities are generated from the last process (arrow) in the graph. B) New metadata model showing that a link to the Sequencing protocol is generated from the ultimate process (process_0) in the graph, and a link to the Library preparation protocol is generated from the penultimate process (process_1) in the graph. From e189bf759857572ecf8dcdeeb953fb4821b7be72 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:16:57 +0100 Subject: [PATCH 03/36] Fixed formatting of things. --- rfcs/text/0000-rfc-library-preparation.md | 47 +++++++++++++++-------- 1 file changed, 32 insertions(+), 15 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 727079ff..f0d001e8 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -45,8 +45,9 @@ The current HCA DCP metadata standard explicitly represents cell suspensions: si From a cell suspension, one (Fig. 1A) or more (Fig. 1B) libraries can be prepared and sequenced. Each library preparation contains cDNA molecules representing a distinct and non-overlapping set of cells. The same library preparation can be sequenced more than once (Fig. 1C), each time producing a unique set of data files. For example, a library preparation can be sequenced on multiple flowcell lanes or a library preparation can be re-sequenced at a later time to generate more data. Regardless of how many times a library preparation was sequenced, **all of the sequence data files derived from one library preparation represent the same set of cells and therefore must be processed together**. Stated another way, **the logical unit of data should be based on a library preparation, all of the sequence files that come from it, and all of the biomaterials, protocols, and processes that generated it**. +--- -![Figure 1](../images/0000-lib_prep_rfc_fig1.png) +![Figure 1](/rfcs/images/0000-lib_prep_rfc_fig1.png) **Figure 1**: Possible droplet-based sequencing experimental designs. A) An experiment where one library preparation was made from a cell suspension and then sequenced once. B) An experiment where two library preparations were made from the same cell suspension and then each library preparation was sequenced once. C) An experiment where two library preparations were made from the same cell suspension and then each library preparation was sequenced twice. Red boxes indicate the set of data files that need to be processed together. @@ -54,6 +55,8 @@ From a cell suspension, one (Fig. 1A) or more (Fig. 1B) libraries can be prepare [Slides](https://drive.google.com/open?id=1vyw6N7qn24qBFAMoKL3nXLHcqpqYFq3Y) prepared by Nick Barkus (particularly slides 13-15) from the June 2019 DCP F2F describe why all sequence data files derived from one library preparation must be processed together. Briefly, a library preparation starts with a set of UMI barcodes attached to transcripts (1 barcode per transcript) and then everything gets amplified (potentially unevenly) and sequenced (Fig. 2A). Sequenced reads are potentially split between different sets of files if the library preparation is sequenced more than once. During processing, unique UMI barcodes are collapsed - meaning identical copies are only counted once - to reflect the original count (Fig. 2B). If files from the same library preparation are processed separately, a collapsed UMI barcode might appear in each set of files, thus inflating the count and leading to the wrong original count (Fig. 2C). +--- + ![Figure 2](/rfcs/images/0000-lib_prep_rfc_fig2.png) **Figure 2**: Processing data files separately inflates UMI counts. A) Two transcripts - each with a unique UMI barcode - for gene A are amplified during library preparation, and the library preparation is sequenced twice to produce two sets of files. B) Files are processed together, and UMI barcodes are collapsed to produce a count of 2 for gene A. C) Files are processed separately, and UMI barcodes are collapsed separately to produce a count of 1 for gene A in the first set of files and 2 for gene A in the second set of files, resulting in an overall count of 3 for gene A (overestimation). @@ -68,10 +71,11 @@ The current HCA metadata standard does not represent a library preparation as an When data contributors supply metadata for their sequence files, they can enter a value for the `sequence_file.library_prep_id` field to indicate which sequence files were generated from the same library preparation. This value must be unique within the spreadsheet (and therefore project since 1 spreadsheet = 1 project), but it does not need to be globally unique (unique within the entire HCA DCP). -| FILE NAME -(Required) | INPUT CELL SUSPENSION ID (Required) | LIBRARY PREPARATION ID (Optional) | +--- + +| FILE NAME (Required) | INPUT CELL SUSPENSION ID (Required) | LIBRARY PREPARATION ID (Optional) | |:-|:-|:-| -| sequence_file.file_core.file_name | cell_suspension.biomaterial_core.biomaterial_id | sequence_file.library_prep_id| +| `sequence_file.file_core.file_name` | `cell_suspension.biomaterial_core.biomaterial_id` | `sequence_file.library_prep_id` | | SRR7159837_1.fastq.gz | cell_suspension_1 | library_preparation_1 | | SRR7159837_2.fastq.gz | cell_suspension_1 | library_preparation_1 | | SRR7159838_1.fastq.gz | cell_suspension_1 | library_preparation_1 | @@ -89,10 +93,14 @@ It is important to consider that, currently, if a valid spreadsheet with the met Assuming a data wrangler or data contributor has filled in the `process_id` field such that each process is assigned a unique identifier, anchoring a logical unit (bundle) on the ultimate process in the graph results in two bundles (Fig. 3A) which means that each bundle does not contain all the data files from the same library preparation. Ideally, logical units are anchored on the library preparation (Fig. 3B) such that all data files from the same library preparation are logically grouped together. -![Figure 3](images/0000-lib_prep_rfc_fig3.png) +--- + +![Figure 3](/rfcs/images/0000-lib_prep_rfc_fig3.png) **Figure 3**: Current and ideal grouping of logical units. A) Logical unit is anchored on ultimate process in the experimental graph, resulting in two bundles which each contain a subset of the data files produced from the same library preparation (not ideal). B) Logical unit is anchored on the library preparation entity, resulting in one logical unit that contains all the data files produced from the same library preparation (ideal). +--- + #### Data consumer perspective Converting the first row from Table 1 (and additional sequence file metadata for that row) to JSON produces: @@ -144,17 +152,19 @@ We will resolve the challenges described above by **creating a first-class libra For cellular resolution experiments, the library preparation entity will have a cell suspension entity as input and sequence file entities as outputs. For bulk cell experiments, the library preparation entity will have a specimen (or organoid or cell line) entity as input and sequence file entities as outputs. Figure 3 below shows the current metadata model representing a droplet-based experiment sequencing a single cell suspension to produce two sets of two files (Fig. 4A). From this experimental design, it is not known whether the two sets of files represent one logical unit or two logical units. If the sequence files were produced from the same library preparation (Fig. 4B), then they represent one logical unit and must be processed together. If the sequence files were produced from different library preparation (Fig. 4C), then they represent two logical units and must be processed separately. -![Figure 4](images/0000-lib_prep_rfc_fig4.png) +--- -**Figure 4**: Determining logical units from droplet-based experimental designs. A) An experiment modeled using the current metadata model which depicts four sequence files derived from one cell suspension. It is unclear what the logical units are. B) An experiment modeled using the proposed metadata model which depicts four sequence files derived from one library preparation. It is clear that these four files represent one logical unit (red outline). C) An experiment modeled using the proposed metadata model which depicts four sequence files derived from two library preparation. It is clear that these four files represent two logical units (one red outline, one blue outline). +![Figure 4](/rfcs/images/0000-lib_prep_rfc_fig4.png) -Arrows in graphs represent processes. Note that in B), two processes were used to derive the two sets of files. Both processes belong to this one logical unit. +**Figure 4**: Determining logical units from droplet-based experimental designs. A) An experiment modeled using the current metadata model which depicts four sequence files derived from one cell suspension. It is unclear what the logical units are. B) An experiment modeled using the proposed metadata model which depicts four sequence files derived from one library preparation. It is clear that these four files represent one logical unit (red outline). C) An experiment modeled using the proposed metadata model which depicts four sequence files derived from two library preparation. It is clear that these four files represent two logical units (one red outline, one blue outline). Arrows in graphs represent processes. Note that in B), two processes were used to derive the two sets of files. Both processes belong to this one logical unit. --- Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided. -![Figure 5](images/0000-lib_prep_rfc_fig5.png) +--- + +![Figure 5](/rfcs/images/0000-lib_prep_rfc_fig5.png) **Figure 5**: Determining logical units from plate-based experimental designs. A) An experiment modeled using the current metadata model which depicts six sequence files derived from three single cell suspension. Logical units are indicated (red, blue, and magenta outlines). B) An experiment modeled using the proposed metadata model which depicts six sequence files derived from three single cell library preparations. Logical units are indicated (red, blue, and magenta outlines). Arrows in graphs represent processes. @@ -213,13 +223,14 @@ Where the `biomaterial_core.json` schema contains the following properties: "supplementary_files": { "description": "A list of filenames of biomaterial-level supplementary files.", }, -"genotype": { +"genotype*": { "description": "Genotype of the biomaterial.", }, -"HDBR_accession": { +"HDBR_accession*": { "description": "A Human Developmental Biology Resource (HDBR) sample accession.", } ``` +> *Consider removing `genotype` and `HDBR_accession` from core as they don't make sense here and in other biomaterials. And the `provenance.json` schema contains the following properties (all supplied by Ingestion Service): @@ -264,10 +275,11 @@ Adding a new first-class biomaterial entity has the potential to add a lot of co Data contributors will supply a project-wide unique ID for each library preparation in the Sequence file tab using the `library_preparation.biomaterial_core.biomaterial_id` field in place of the current `sequence_file.library_prep_id` field. When the spreadsheet is imported, the Ingestion Service will generate a globally unique UUID for each library preparation entity. Unlike other biomaterials, there would not be a separate spreadsheet tab for library preparation entities for two reasons. 1. Adding a spreadsheet tab adds considerable complexity for data contributors. 2. There are only two fields that need to be supplied by the data contributor for which generating a whole new tab creates high overhead, especially considering one of the fields replaces a field already in the Sequence file tab. Without the ability to automatically infer `library_preparation.biomaterial_core.ncbi_taxon_id`, this field would need to be added to the Sequence file tab and would need to be filled in by contributors. For example: +--- | FILE NAME (Required) | INPUT CELL SUSPENSION ID (Required) | INPUT LIBRARY PREPARATION ID (Required) | NCBI TAXON ID (Required) | |:-|:-|:-|:-| -| sequence_file.file_core.file_name | cell_suspension.biomaterial_core.biomaterial_id | library_preparation.biomaterial_core.biomaterial_id | library_preparation.biomaterial_core.ncbi_taxon_id | +| `sequence_file.file_core.file_name` | `cell_suspension.biomaterial_core.biomaterial_id` | `library_preparation.biomaterial_core.biomaterial_id` | `library_preparation.biomaterial_core.ncbi_taxon_id` | | SRR7159837_1.fastq.gz | cell_suspension_1 | library_preparation_1 | 9606 | | SRR7159837_2.fastq.gz | cell_suspension_1 | library_preparation_1 | 9606 | | SRR7159838_1.fastq.gz | cell_suspension_1 | library_preparation_2 | 9606 | @@ -310,10 +322,11 @@ Data consumers will benefit from the metadata model now aligning with INSDC “e - Library preparation is equivalent to an INSDC experiment - Single set of files from a library preparation is equivalent to an INSDC run +--- | FILE NAME | INPUT LIBRARY PREPARATION ID | INSDC EXPERIMENT ACCESSION | INSDC RUN ACCESSIONS | |:-|:-|:-|:-| -| sequence_file.file_core.file_name | library_preparation.biomaterial_core.biomaterial_id | process.insdc_experiment.insdc_experiment_accession | sequence_file.insdc_run_accessions | +| `sequence_file.file_core.file_name` | `library_preparation.biomaterial_core.biomaterial_id` | `process.insdc_experiment.insdc_experiment_accession` | `sequence_file.insdc_run_accessions` | | SRR7159837_1.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159837 | | SRR7159837_2.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159837 | | SRR7159838_1.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159838 | @@ -335,9 +348,13 @@ Data consumers will benefit from the metadata model now aligning with INSDC “e - Process_0 <-> Sequencing protocol - Declare rules about linking -![Figure 6](images/0000-lib_prep_rfc_fig6.png) +--- + +![Figure 6](/rfcs/images/0000-lib_prep_rfc_fig6.png) -Figure 5: Diagram of how protocol linking will change with new library preparation entity. A) Current metadata model showing that links to Sequencing protocol and Library preparation protocol entities are generated from the last process (arrow) in the graph. B) New metadata model showing that a link to the Sequencing protocol is generated from the ultimate process (process_0) in the graph, and a link to the Library preparation protocol is generated from the penultimate process (process_1) in the graph. +Figure 6: Diagram of how protocol linking will change with new library preparation entity. A) Current metadata model showing that links to Sequencing protocol and Library preparation protocol entities are generated from the last process (arrow) in the graph. B) New metadata model showing that a link to the Sequencing protocol is generated from the ultimate process (process_0) in the graph, and a link to the Library preparation protocol is generated from the penultimate process (process_1) in the graph. + +--- #### Data Storage Service From e514cd44062feaffdbd0f08c2ef56fde7a655bf2 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:19:41 +0100 Subject: [PATCH 04/36] Fixed formatting of things. --- rfcs/text/0000-rfc-library-preparation.md | 14 ++++---------- 1 file changed, 4 insertions(+), 10 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index f0d001e8..4f05b9ab 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -53,7 +53,7 @@ From a cell suspension, one (Fig. 1A) or more (Fig. 1B) libraries can be prepare --- -[Slides](https://drive.google.com/open?id=1vyw6N7qn24qBFAMoKL3nXLHcqpqYFq3Y) prepared by Nick Barkus (particularly slides 13-15) from the June 2019 DCP F2F describe why all sequence data files derived from one library preparation must be processed together. Briefly, a library preparation starts with a set of UMI barcodes attached to transcripts (1 barcode per transcript) and then everything gets amplified (potentially unevenly) and sequenced (Fig. 2A). Sequenced reads are potentially split between different sets of files if the library preparation is sequenced more than once. During processing, unique UMI barcodes are collapsed - meaning identical copies are only counted once - to reflect the original count (Fig. 2B). If files from the same library preparation are processed separately, a collapsed UMI barcode might appear in each set of files, thus inflating the count and leading to the wrong original count (Fig. 2C). +[Slides](https://drive.google.com/open?id=1vyw6N7qn24qBFAMoKL3nXLHcqpqYFq3Y) prepared by Nick Barkus (particularly slides 13-15) from the June 2019 DCP F2F describe why all sequence data files derived from one library preparation must be processed together. Briefly, a library preparation starts with a set of UMI barcodes attached to transcripts (1 barcode per transcript) and then everything gets amplified (potentially unevenly) and sequenced (Fig. 2A). Reads are potentially split between different sets of files if the library preparation is sequenced more than once. During processing, unique UMI barcodes are collapsed - identical copies are only counted once - to calculate the original count (Fig. 2B). If files from the same library preparation are processed separately, a collapsed UMI barcode might appear in each set of files, thus inflating the count and leading to the wrong original count (Fig. 2C). --- @@ -71,11 +71,9 @@ The current HCA metadata standard does not represent a library preparation as an When data contributors supply metadata for their sequence files, they can enter a value for the `sequence_file.library_prep_id` field to indicate which sequence files were generated from the same library preparation. This value must be unique within the spreadsheet (and therefore project since 1 spreadsheet = 1 project), but it does not need to be globally unique (unique within the entire HCA DCP). ---- - | FILE NAME (Required) | INPUT CELL SUSPENSION ID (Required) | LIBRARY PREPARATION ID (Optional) | |:-|:-|:-| -| `sequence_file.file_core.file_name` | `cell_suspension.biomaterial_core.biomaterial_id` | `sequence_file.library_prep_id` | +| `sequence_file.file_core. file_name` | `cell_suspension.biomaterial_core. biomaterial_id` | `sequence_file.library_prep_id` | | SRR7159837_1.fastq.gz | cell_suspension_1 | library_preparation_1 | | SRR7159837_2.fastq.gz | cell_suspension_1 | library_preparation_1 | | SRR7159838_1.fastq.gz | cell_suspension_1 | library_preparation_1 | @@ -274,12 +272,10 @@ The required fields for the library preparation entity will be: Adding a new first-class biomaterial entity has the potential to add a lot of complexity to the metadata model. One area we want to avoid adding complexity is in the metadata spreadsheet, which is already quite complex for data contributors to fill out. Given that there are only two required (user-supplied) metadata fields proposed for library preparation entities, it is possible to keep the metadata spreadsheet relatively unchanged. Data contributors will supply a project-wide unique ID for each library preparation in the Sequence file tab using the `library_preparation.biomaterial_core.biomaterial_id` field in place of the current `sequence_file.library_prep_id` field. When the spreadsheet is imported, the Ingestion Service will generate a globally unique UUID for each library preparation entity. Unlike other biomaterials, there would not be a separate spreadsheet tab for library preparation entities for two reasons. 1. Adding a spreadsheet tab adds considerable complexity for data contributors. 2. There are only two fields that need to be supplied by the data contributor for which generating a whole new tab creates high overhead, especially considering one of the fields replaces a field already in the Sequence file tab. Without the ability to automatically infer `library_preparation.biomaterial_core.ncbi_taxon_id`, this field would need to be added to the Sequence file tab and would need to be filled in by contributors. For example: - ---- | FILE NAME (Required) | INPUT CELL SUSPENSION ID (Required) | INPUT LIBRARY PREPARATION ID (Required) | NCBI TAXON ID (Required) | |:-|:-|:-|:-| -| `sequence_file.file_core.file_name` | `cell_suspension.biomaterial_core.biomaterial_id` | `library_preparation.biomaterial_core.biomaterial_id` | `library_preparation.biomaterial_core.ncbi_taxon_id` | +| `sequence_file.file_core. file_name` | `cell_suspension.biomaterial_core. biomaterial_id` | `library_preparation.biomaterial_core. biomaterial_id` | `library_preparation.biomaterial_core. ncbi_taxon_id` | | SRR7159837_1.fastq.gz | cell_suspension_1 | library_preparation_1 | 9606 | | SRR7159837_2.fastq.gz | cell_suspension_1 | library_preparation_1 | 9606 | | SRR7159838_1.fastq.gz | cell_suspension_1 | library_preparation_2 | 9606 | @@ -322,11 +318,9 @@ Data consumers will benefit from the metadata model now aligning with INSDC “e - Library preparation is equivalent to an INSDC experiment - Single set of files from a library preparation is equivalent to an INSDC run ---- - | FILE NAME | INPUT LIBRARY PREPARATION ID | INSDC EXPERIMENT ACCESSION | INSDC RUN ACCESSIONS | |:-|:-|:-|:-| -| `sequence_file.file_core.file_name` | `library_preparation.biomaterial_core.biomaterial_id` | `process.insdc_experiment.insdc_experiment_accession` | `sequence_file.insdc_run_accessions` | +| `sequence_file.file_core. file_name` | `library_preparation.biomaterial_core. biomaterial_id` | `process.insdc_experiment. insdc_experiment_accession` | `sequence_file. insdc_run_accessions` | | SRR7159837_1.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159837 | | SRR7159837_2.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159837 | | SRR7159838_1.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159838 | From ad190ca0133a0a20662e3e883d294b53a972f716 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:25:06 +0100 Subject: [PATCH 05/36] Tested resizing image. --- rfcs/text/0000-rfc-library-preparation.md | 2 ++ 1 file changed, 2 insertions(+) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 4f05b9ab..7609ab3a 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -95,6 +95,8 @@ Assuming a data wrangler or data contributor has filled in the `process_id` fiel ![Figure 3](/rfcs/images/0000-lib_prep_rfc_fig3.png) + + **Figure 3**: Current and ideal grouping of logical units. A) Logical unit is anchored on ultimate process in the experimental graph, resulting in two bundles which each contain a subset of the data files produced from the same library preparation (not ideal). B) Logical unit is anchored on the library preparation entity, resulting in one logical unit that contains all the data files produced from the same library preparation (ideal). --- From eab833ca5ffa57fea8315758988a9cda55ffabc1 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:25:40 +0100 Subject: [PATCH 06/36] Tested resizing image. --- rfcs/text/0000-rfc-library-preparation.md | 4 +--- 1 file changed, 1 insertion(+), 3 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 7609ab3a..b177d6aa 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -93,9 +93,7 @@ Assuming a data wrangler or data contributor has filled in the `process_id` fiel --- -![Figure 3](/rfcs/images/0000-lib_prep_rfc_fig3.png) - - + **Figure 3**: Current and ideal grouping of logical units. A) Logical unit is anchored on ultimate process in the experimental graph, resulting in two bundles which each contain a subset of the data files produced from the same library preparation (not ideal). B) Logical unit is anchored on the library preparation entity, resulting in one logical unit that contains all the data files produced from the same library preparation (ideal). From 7489fbb57bccefc93e6a5b2d74fc48a6da46e717 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:26:25 +0100 Subject: [PATCH 07/36] Tested resizing image. --- rfcs/text/0000-rfc-library-preparation.md | 6 +++--- 1 file changed, 3 insertions(+), 3 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index b177d6aa..8df9bf1d 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -93,7 +93,7 @@ Assuming a data wrangler or data contributor has filled in the `process_id` fiel --- - + **Figure 3**: Current and ideal grouping of logical units. A) Logical unit is anchored on ultimate process in the experimental graph, resulting in two bundles which each contain a subset of the data files produced from the same library preparation (not ideal). B) Logical unit is anchored on the library preparation entity, resulting in one logical unit that contains all the data files produced from the same library preparation (ideal). @@ -152,7 +152,7 @@ For cellular resolution experiments, the library preparation entity will have a --- -![Figure 4](/rfcs/images/0000-lib_prep_rfc_fig4.png) + **Figure 4**: Determining logical units from droplet-based experimental designs. A) An experiment modeled using the current metadata model which depicts four sequence files derived from one cell suspension. It is unclear what the logical units are. B) An experiment modeled using the proposed metadata model which depicts four sequence files derived from one library preparation. It is clear that these four files represent one logical unit (red outline). C) An experiment modeled using the proposed metadata model which depicts four sequence files derived from two library preparation. It is clear that these four files represent two logical units (one red outline, one blue outline). Arrows in graphs represent processes. Note that in B), two processes were used to derive the two sets of files. Both processes belong to this one logical unit. @@ -162,7 +162,7 @@ Figure 5 below shows the current metadata model representing a plate-based exper --- -![Figure 5](/rfcs/images/0000-lib_prep_rfc_fig5.png) + **Figure 5**: Determining logical units from plate-based experimental designs. A) An experiment modeled using the current metadata model which depicts six sequence files derived from three single cell suspension. Logical units are indicated (red, blue, and magenta outlines). B) An experiment modeled using the proposed metadata model which depicts six sequence files derived from three single cell library preparations. Logical units are indicated (red, blue, and magenta outlines). Arrows in graphs represent processes. From 2cbc221fa9451d13716f58eb54189ffc853ded7e Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:28:11 +0100 Subject: [PATCH 08/36] Tested resizing image. --- rfcs/text/0000-rfc-library-preparation.md | 8 ++++---- 1 file changed, 4 insertions(+), 4 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 8df9bf1d..27733381 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -337,16 +337,16 @@ Data consumers will benefit from the metadata model now aligning with INSDC “e - Logical units will be anchored on a library preparation entity UUID (or possibly the process entity that is input to the library preparation entity) in the experimental graph (exporter). - Schema template generator will be modified to (1) put the library preparation entity fields in the Sequence file tab, and (2) exclude all the library preparation entity fields except the required ones. - Experimental graph (instantiated by links.json) will represent that (1) a library preparation protocol is followed to produce a library preparation from a cell suspension (or other biomaterial for bulk sequencing) and that (2) a sequencing protocol is followed to produce sequence file(s) from a library preparation. Users of the ingest-api will supply these links during programmatic submission. Otherwise, Ingestion Service (ingest-broker) will create the following links from metadata in the Sequence file tab and some graph assumptions (Fig. 6): - - Cell suspension <-> Process_1 <-> Library preparation <-> Process_0 <-> Sequence file - - Process_1 <-> Library preparation protocol - - Process_0 <-> Sequencing protocol + - Cell suspension :left_right_arrow: Process_1 :left_right_arrow: Library preparation :left_right_arrow: Process_0 :left_right_arrow: Sequence file + - Process_1 :left_right_arrow: Library preparation protocol + - Process_0 :left_right_arrow: Sequencing protocol - Declare rules about linking --- ![Figure 6](/rfcs/images/0000-lib_prep_rfc_fig6.png) -Figure 6: Diagram of how protocol linking will change with new library preparation entity. A) Current metadata model showing that links to Sequencing protocol and Library preparation protocol entities are generated from the last process (arrow) in the graph. B) New metadata model showing that a link to the Sequencing protocol is generated from the ultimate process (process_0) in the graph, and a link to the Library preparation protocol is generated from the penultimate process (process_1) in the graph. +**Figure 6**: Diagram of how protocol linking will change with new library preparation entity. A) Current metadata model showing that links to Sequencing protocol and Library preparation protocol entities are generated from the last process (arrow) in the graph. B) New metadata model showing that a link to the Sequencing protocol is generated from the ultimate process (process_0) in the graph, and a link to the Library preparation protocol is generated from the penultimate process (process_1) in the graph. --- From cdfd5339696c0d11487ecef859edb65561751ea0 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:31:37 +0100 Subject: [PATCH 09/36] Tested resizing image. --- rfcs/text/0000-rfc-library-preparation.md | 8 ++++++-- 1 file changed, 6 insertions(+), 2 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 27733381..c65420cc 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -96,6 +96,9 @@ Assuming a data wrangler or data contributor has filled in the `process_id` fiel **Figure 3**: Current and ideal grouping of logical units. A) Logical unit is anchored on ultimate process in the experimental graph, resulting in two bundles which each contain a subset of the data files produced from the same library preparation (not ideal). B) Logical unit is anchored on the library preparation entity, resulting in one logical unit that contains all the data files produced from the same library preparation (ideal). +. +. +. --- @@ -152,17 +155,18 @@ For cellular resolution experiments, the library preparation entity will have a --- - + **Figure 4**: Determining logical units from droplet-based experimental designs. A) An experiment modeled using the current metadata model which depicts four sequence files derived from one cell suspension. It is unclear what the logical units are. B) An experiment modeled using the proposed metadata model which depicts four sequence files derived from one library preparation. It is clear that these four files represent one logical unit (red outline). C) An experiment modeled using the proposed metadata model which depicts four sequence files derived from two library preparation. It is clear that these four files represent two logical units (one red outline, one blue outline). Arrows in graphs represent processes. Note that in B), two processes were used to derive the two sets of files. Both processes belong to this one logical unit. --- + Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided. --- - + **Figure 5**: Determining logical units from plate-based experimental designs. A) An experiment modeled using the current metadata model which depicts six sequence files derived from three single cell suspension. Logical units are indicated (red, blue, and magenta outlines). B) An experiment modeled using the proposed metadata model which depicts six sequence files derived from three single cell library preparations. Logical units are indicated (red, blue, and magenta outlines). Arrows in graphs represent processes. From 8c599811f0de002ef0e9b922c2319be2080d1b37 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:35:10 +0100 Subject: [PATCH 10/36] Fixed formatting of things. --- rfcs/text/0000-rfc-library-preparation.md | 13 +++++++------ 1 file changed, 7 insertions(+), 6 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index c65420cc..13ac6dcc 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -95,10 +95,11 @@ Assuming a data wrangler or data contributor has filled in the `process_id` fiel -**Figure 3**: Current and ideal grouping of logical units. A) Logical unit is anchored on ultimate process in the experimental graph, resulting in two bundles which each contain a subset of the data files produced from the same library preparation (not ideal). B) Logical unit is anchored on the library preparation entity, resulting in one logical unit that contains all the data files produced from the same library preparation (ideal). -. -. -. +**Figure 3**: Current and ideal grouping of logical units. A) Logical unit is anchored on ultimate process in the experimental graph, resulting in two bundles which each contain a subset of the data files produced from the same library preparation (not ideal). B) Logical unit is anchored on the library preparation entity, resulting in one logical unit that contains all the data files produced from the same library preparation (ideal).\ +
+
+
+
--- @@ -157,7 +158,7 @@ For cellular resolution experiments, the library preparation entity will have a -**Figure 4**: Determining logical units from droplet-based experimental designs. A) An experiment modeled using the current metadata model which depicts four sequence files derived from one cell suspension. It is unclear what the logical units are. B) An experiment modeled using the proposed metadata model which depicts four sequence files derived from one library preparation. It is clear that these four files represent one logical unit (red outline). C) An experiment modeled using the proposed metadata model which depicts four sequence files derived from two library preparation. It is clear that these four files represent two logical units (one red outline, one blue outline). Arrows in graphs represent processes. Note that in B), two processes were used to derive the two sets of files. Both processes belong to this one logical unit. +**Figure 4: Determining logical units from droplet-based experimental designs**. A) An experiment modeled using the current metadata model which depicts four sequence files derived from one cell suspension. It is unclear what the logical units are. B) An experiment modeled using the proposed metadata model which depicts four sequence files derived from one library preparation. It is clear that the four files represent one logical unit (red outline). C) An experiment modeled using the proposed metadata model which depicts four sequence files derived from two library preparations. It is clear that the four files represent two logical units. Arrows in graphs represent processes. Note that in B), two processes were used to derive the two sets of files. Both processes belong to this one logical unit. --- @@ -166,7 +167,7 @@ Figure 5 below shows the current metadata model representing a plate-based exper --- - + **Figure 5**: Determining logical units from plate-based experimental designs. A) An experiment modeled using the current metadata model which depicts six sequence files derived from three single cell suspension. Logical units are indicated (red, blue, and magenta outlines). B) An experiment modeled using the proposed metadata model which depicts six sequence files derived from three single cell library preparations. Logical units are indicated (red, blue, and magenta outlines). Arrows in graphs represent processes. From 36de394a437c8b09281ee3cc7f92eaa45728a363 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:35:58 +0100 Subject: [PATCH 11/36] Fixed formatting of things. --- rfcs/text/0000-rfc-library-preparation.md | 10 +++++----- 1 file changed, 5 insertions(+), 5 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 13ac6dcc..ceae262d 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -49,7 +49,7 @@ From a cell suspension, one (Fig. 1A) or more (Fig. 1B) libraries can be prepare ![Figure 1](/rfcs/images/0000-lib_prep_rfc_fig1.png) -**Figure 1**: Possible droplet-based sequencing experimental designs. A) An experiment where one library preparation was made from a cell suspension and then sequenced once. B) An experiment where two library preparations were made from the same cell suspension and then each library preparation was sequenced once. C) An experiment where two library preparations were made from the same cell suspension and then each library preparation was sequenced twice. Red boxes indicate the set of data files that need to be processed together. +**Figure 1: Possible droplet-based sequencing experimental designs**. A) An experiment where one library preparation was made from a cell suspension and then sequenced once. B) An experiment where two library preparations were made from the same cell suspension and then each library preparation was sequenced once. C) An experiment where two library preparations were made from the same cell suspension and then each library preparation was sequenced twice. Red boxes indicate the set of data files that need to be processed together. --- @@ -59,7 +59,7 @@ From a cell suspension, one (Fig. 1A) or more (Fig. 1B) libraries can be prepare ![Figure 2](/rfcs/images/0000-lib_prep_rfc_fig2.png) -**Figure 2**: Processing data files separately inflates UMI counts. A) Two transcripts - each with a unique UMI barcode - for gene A are amplified during library preparation, and the library preparation is sequenced twice to produce two sets of files. B) Files are processed together, and UMI barcodes are collapsed to produce a count of 2 for gene A. C) Files are processed separately, and UMI barcodes are collapsed separately to produce a count of 1 for gene A in the first set of files and 2 for gene A in the second set of files, resulting in an overall count of 3 for gene A (overestimation). +**Figure 2: Processing data files separately inflates UMI counts**. A) Two transcripts - each with a unique UMI barcode - for gene A are amplified during library preparation, and the library preparation is sequenced twice to produce two sets of files. B) Files are processed together, and UMI barcodes are collapsed to produce a count of 2 for gene A. C) Files are processed separately, and UMI barcodes are collapsed separately to produce a count of 1 for gene A in the first set of files and 2 for gene A in the second set of files, resulting in an overall count of 3 for gene A (overestimation). --- @@ -95,7 +95,7 @@ Assuming a data wrangler or data contributor has filled in the `process_id` fiel -**Figure 3**: Current and ideal grouping of logical units. A) Logical unit is anchored on ultimate process in the experimental graph, resulting in two bundles which each contain a subset of the data files produced from the same library preparation (not ideal). B) Logical unit is anchored on the library preparation entity, resulting in one logical unit that contains all the data files produced from the same library preparation (ideal).\ +**Figure 3: Current and ideal grouping of logical units**. A) Logical unit is anchored on ultimate process in the experimental graph, resulting in two bundles which each contain a subset of the data files produced from the same library preparation (not ideal). B) Logical unit is anchored on the library preparation entity, resulting in one logical unit that contains all the data files produced from the same library preparation (ideal).\


@@ -169,7 +169,7 @@ Figure 5 below shows the current metadata model representing a plate-based exper -**Figure 5**: Determining logical units from plate-based experimental designs. A) An experiment modeled using the current metadata model which depicts six sequence files derived from three single cell suspension. Logical units are indicated (red, blue, and magenta outlines). B) An experiment modeled using the proposed metadata model which depicts six sequence files derived from three single cell library preparations. Logical units are indicated (red, blue, and magenta outlines). Arrows in graphs represent processes. +**Figure 5: Determining logical units from plate-based experimental designs**. A) An experiment modeled using the current metadata model which depicts six sequence files derived from three single cell suspension. Logical units are indicated (red, blue, and magenta outlines). B) An experiment modeled using the proposed metadata model which depicts six sequence files derived from three single cell library preparations. Logical units are indicated (red, blue, and magenta outlines). Arrows in graphs represent processes. **NB**: An experimental design where the same library preparation is sequenced more than once is not shown here. This is either (1) not even feasible or (2) highly unlikely to occur given that there isn’t a lot of material to sequence the same cell more than once. @@ -351,7 +351,7 @@ Data consumers will benefit from the metadata model now aligning with INSDC “e ![Figure 6](/rfcs/images/0000-lib_prep_rfc_fig6.png) -**Figure 6**: Diagram of how protocol linking will change with new library preparation entity. A) Current metadata model showing that links to Sequencing protocol and Library preparation protocol entities are generated from the last process (arrow) in the graph. B) New metadata model showing that a link to the Sequencing protocol is generated from the ultimate process (process_0) in the graph, and a link to the Library preparation protocol is generated from the penultimate process (process_1) in the graph. +**Figure 6: Diagram of how protocol linking will change with new library preparation entity**. A) Current metadata model showing that links to Sequencing protocol and Library preparation protocol entities are generated from the last process (arrow) in the graph. B) New metadata model showing that a link to the Sequencing protocol is generated from the ultimate process (process_0) in the graph, and a link to the Library preparation protocol is generated from the penultimate process (process_1) in the graph. --- From b8fb931c0a18a0b2a15d39e23e1ffcbf41dbee8c Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:36:56 +0100 Subject: [PATCH 12/36] Fixed formatting of things. --- rfcs/text/0000-rfc-library-preparation.md | 1 - 1 file changed, 1 deletion(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index ceae262d..06269bbd 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -162,7 +162,6 @@ For cellular resolution experiments, the library preparation entity will have a --- - Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided. --- From 21bc457cffced1f49ae8cb82a3cfe3109d1ee1db Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:37:39 +0100 Subject: [PATCH 13/36] Debugging. --- rfcs/text/0000-rfc-library-preparation.md | 2 ++ 1 file changed, 2 insertions(+) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 06269bbd..911fdcc8 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -162,6 +162,8 @@ For cellular resolution experiments, the library preparation entity will have a --- +What is going on. + Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided. --- From 7f23ebaf2054c238118bdf214529fdfe23999b8a Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:38:04 +0100 Subject: [PATCH 14/36] Debugging. --- rfcs/text/0000-rfc-library-preparation.md | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 911fdcc8..5e3f5517 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -162,9 +162,9 @@ For cellular resolution experiments, the library preparation entity will have a --- -What is going on. +What is going on. Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided. -Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided. +What is going on. --- From 05dcbab8d0ac8fde45dd11c1273e353486e98cb6 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:38:25 +0100 Subject: [PATCH 15/36] Debugging. --- rfcs/text/0000-rfc-library-preparation.md | 2 -- 1 file changed, 2 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 5e3f5517..aa2ce0f3 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -163,8 +163,6 @@ For cellular resolution experiments, the library preparation entity will have a --- What is going on. Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided. - -What is going on. --- From 17db6f89b1c804901cb2822473067d570e8cce62 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:38:56 +0100 Subject: [PATCH 16/36] Debugging. --- rfcs/text/0000-rfc-library-preparation.md | 4 +++- 1 file changed, 3 insertions(+), 1 deletion(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index aa2ce0f3..547095e9 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -162,7 +162,9 @@ For cellular resolution experiments, the library preparation entity will have a --- -What is going on. Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided. +Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided.\ + +. --- From 42bfd2604346b343c4510351881b702467a78d48 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:39:30 +0100 Subject: [PATCH 17/36] Debugging. --- rfcs/text/0000-rfc-library-preparation.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 547095e9..b0f31a6c 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -162,7 +162,7 @@ For cellular resolution experiments, the library preparation entity will have a --- -Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided.\ +Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided.
. From 40cc466235847cf2a24b6d7e4c3550ed1507ddb2 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:44:11 +0100 Subject: [PATCH 18/36] Debugging. --- rfcs/text/0000-rfc-library-preparation.md | 27 +++++++++++------------ 1 file changed, 13 insertions(+), 14 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index b0f31a6c..78bdd666 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -163,9 +163,8 @@ For cellular resolution experiments, the library preparation entity will have a --- Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided.
- . - + --- @@ -279,19 +278,19 @@ Adding a new first-class biomaterial entity has the potential to add a lot of co Data contributors will supply a project-wide unique ID for each library preparation in the Sequence file tab using the `library_preparation.biomaterial_core.biomaterial_id` field in place of the current `sequence_file.library_prep_id` field. When the spreadsheet is imported, the Ingestion Service will generate a globally unique UUID for each library preparation entity. Unlike other biomaterials, there would not be a separate spreadsheet tab for library preparation entities for two reasons. 1. Adding a spreadsheet tab adds considerable complexity for data contributors. 2. There are only two fields that need to be supplied by the data contributor for which generating a whole new tab creates high overhead, especially considering one of the fields replaces a field already in the Sequence file tab. Without the ability to automatically infer `library_preparation.biomaterial_core.ncbi_taxon_id`, this field would need to be added to the Sequence file tab and would need to be filled in by contributors. For example: -| FILE NAME (Required) | INPUT CELL SUSPENSION ID (Required) | INPUT LIBRARY PREPARATION ID (Required) | NCBI TAXON ID (Required) | +| FILE NAME (Required) | *INPUT* CELL SUSPENSION ID (*Required*) | INPUT LIBRARY PREPARATION ID (Required) | *NCBI TAXON ID (Required)* | |:-|:-|:-|:-| -| `sequence_file.file_core. file_name` | `cell_suspension.biomaterial_core. biomaterial_id` | `library_preparation.biomaterial_core. biomaterial_id` | `library_preparation.biomaterial_core. ncbi_taxon_id` | -| SRR7159837_1.fastq.gz | cell_suspension_1 | library_preparation_1 | 9606 | -| SRR7159837_2.fastq.gz | cell_suspension_1 | library_preparation_1 | 9606 | -| SRR7159838_1.fastq.gz | cell_suspension_1 | library_preparation_2 | 9606 | -| SRR7159838_2.fastq.gz | cell_suspension_1 | library_preparation_2 | 9606 | -| SRR7159839_1.fastq.gz | cell_suspension_1 | library_preparation_3 | 9606 | -| SRR7159839_2.fastq.gz | cell_suspension_1 | library_preparation_3 | 9606 | -| SRR7159840_1.fastq.gz | cell_suspension_1 | library_preparation_4 | 9606 | -| SRR7159840_2.fastq.gz | cell_suspension_1 | library_preparation_4 | 9606 | - -**Table 2**: An example set of sequence file metadata based on Fig. 1B experimental design. User-friendly field names in row 1; fully-qualified programmatic field names in row 2. Text in red indicates spreadsheet differences between Table 1 and Table 2. +| `sequence_file.file_core. file_name` | `cell_suspension.biomaterial_core. biomaterial_id` | *`library_preparation.biomaterial_core. biomaterial_id`* | *`library_preparation.biomaterial_core. ncbi_taxon_id`* | +| SRR7159837_1.fastq.gz | cell_suspension_1 | library_preparation_1 | *9606* | +| SRR7159837_2.fastq.gz | cell_suspension_1 | library_preparation_1 | *9606* | +| SRR7159838_1.fastq.gz | cell_suspension_1 | library_preparation_2 | *9606* | +| SRR7159838_2.fastq.gz | cell_suspension_1 | library_preparation_2 | *9606* | +| SRR7159839_1.fastq.gz | cell_suspension_1 | library_preparation_3 | *9606* | +| SRR7159839_2.fastq.gz | cell_suspension_1 | library_preparation_3 | *9606* | +| SRR7159840_1.fastq.gz | cell_suspension_1 | library_preparation_4 | *9606* | +| SRR7159840_2.fastq.gz | cell_suspension_1 | library_preparation_4 | *9606* | + +**Table 2**: An example set of sequence file metadata based on Fig. 1B experimental design. User-friendly field names in row 1; fully-qualified programmatic field names in row 2. Italicized text indicates spreadsheet differences between Table 1 and Table 2. --- From 17b0b6c407e767dd08978a90ebe028c855c9a134 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 16:45:26 +0100 Subject: [PATCH 19/36] Debugging. --- rfcs/text/0000-rfc-library-preparation.md | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 78bdd666..7d05d6d6 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -163,7 +163,7 @@ For cellular resolution experiments, the library preparation entity will have a --- Figure 5 below shows the current metadata model representing a plate-based experiment sequencing three single cell suspensions to produce three sets of two files (Fig. 5A). From this experimental design, it is clear that each set of files represents one logical unit (red, blue, and magenta outlines). The experimental design and logical units are equally as clear under the new proposed metadata model (Fig. 5B). For plate-based sequencing, each cell goes through a library construction protocol to produce a library preparation. These library preparations are then pooled, sequenced, and demultiplexed such that per-cell suspension sequence files are provided.
-. + --- @@ -280,7 +280,7 @@ Data contributors will supply a project-wide unique ID for each library preparat | FILE NAME (Required) | *INPUT* CELL SUSPENSION ID (*Required*) | INPUT LIBRARY PREPARATION ID (Required) | *NCBI TAXON ID (Required)* | |:-|:-|:-|:-| -| `sequence_file.file_core. file_name` | `cell_suspension.biomaterial_core. biomaterial_id` | *`library_preparation.biomaterial_core. biomaterial_id`* | *`library_preparation.biomaterial_core. ncbi_taxon_id`* | +| `sequence_file.file_core. file_name` | `cell_suspension.biomaterial_ core.biomaterial_id` | *`library_preparation.biomaterial_ core.biomaterial_id`* | *`library_preparation.biomaterial_ core.ncbi_taxon_id`* | | SRR7159837_1.fastq.gz | cell_suspension_1 | library_preparation_1 | *9606* | | SRR7159837_2.fastq.gz | cell_suspension_1 | library_preparation_1 | *9606* | | SRR7159838_1.fastq.gz | cell_suspension_1 | library_preparation_2 | *9606* | From 8bbcb85630923dae4cf2c4b1e2e46bbde4d95dd1 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 17:01:40 +0100 Subject: [PATCH 20/36] Final formatting fixes. --- rfcs/text/0000-rfc-library-preparation.md | 32 +++++++++++------------ 1 file changed, 15 insertions(+), 17 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 7d05d6d6..764e3811 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -8,9 +8,7 @@ ## Summary -The current HCA DCP metadata model explicitly represents cell suspensions (single cells or multiple cells suspended in some media) but not the sequencing library preparations derived from them. This is creating challenges for contributors, consumers, and DCP implementation teams when submitting, processing, and interpreting sequencing data. - -Here we propose a solution for explicitly identifying library preparation biomaterials in a sequencing experiment by making them a first-class biomaterial type entity in the metadata schema alongside donor, cell suspension, imaged specimen, etc. +The current HCA DCP metadata model explicitly represents cell suspensions (single cells or multiple cells suspended in some media) but not the sequencing library preparations derived from them. This is creating challenges for contributors, consumers, and DCP implementation teams when submitting, processing, and interpreting sequencing data. Here we propose a solution for explicitly identifying library preparation biomaterials in a sequencing experiment by making them a first-class biomaterial type entity in the metadata standard. ## Author(s) @@ -53,7 +51,7 @@ From a cell suspension, one (Fig. 1A) or more (Fig. 1B) libraries can be prepare --- -[Slides](https://drive.google.com/open?id=1vyw6N7qn24qBFAMoKL3nXLHcqpqYFq3Y) prepared by Nick Barkus (particularly slides 13-15) from the June 2019 DCP F2F describe why all sequence data files derived from one library preparation must be processed together. Briefly, a library preparation starts with a set of UMI barcodes attached to transcripts (1 barcode per transcript) and then everything gets amplified (potentially unevenly) and sequenced (Fig. 2A). Reads are potentially split between different sets of files if the library preparation is sequenced more than once. During processing, unique UMI barcodes are collapsed - identical copies are only counted once - to calculate the original count (Fig. 2B). If files from the same library preparation are processed separately, a collapsed UMI barcode might appear in each set of files, thus inflating the count and leading to the wrong original count (Fig. 2C). +[Slides](https://drive.google.com/open?id=1vyw6N7qn24qBFAMoKL3nXLHcqpqYFq3Y) by Nick Barkus (particularly slides 13-15) from the June 2019 DCP F2F describe why all sequence data files derived from one library preparation must be processed together. Briefly, a library preparation starts with a set of UMI barcodes attached to transcripts (1 barcode per transcript), and then everything gets amplified (potentially unevenly) and sequenced (Fig. 2A). Reads are potentially split between different sets of files if the library preparation is sequenced more than once. During processing, unique UMI barcodes are collapsed - identical copies are only counted once - to calculate the original count (Fig. 2B). If files from the same library preparation are processed separately, a collapsed UMI barcode might appear in each set of files, thus inflating the count and leading to an inaccurate original count (Fig. 2C). --- @@ -175,7 +173,7 @@ Figure 5 below shows the current metadata model representing a plate-based exper --- -This proposal will require modifications to some DCP components (see “Implementation changes for DCP components” section below for more details). Importantly, with a new library preparation entity, the logical unit of data should be anchored on the library preparation, not the ultimate process used to generate a sequence file (colloquially referred to as the “assay process” in the Ingestion Service). +This proposal will require modifications to some DCP components (see "Implementation changes for DCP components" section below for more details). Importantly, with a new library preparation entity, the logical unit of data should be anchored on the library preparation, not the ultimate process used to generate a sequence file (colloquially referred to as the "assay process" in the Ingestion Service). ### Metadata schema for library preparation entity @@ -316,22 +314,22 @@ Data consumers will be able to find all data that need to be processed together will return all bundles (primary and secondary) that contain the specified library preparation. Data consumers will not need to depend on multi-bundle notifications for processing all data files from the same library preparation.. All data files from a single library preparation will be put in the same logical unit if submitted at one time (in one submission envelope). Caveats: -1. Still unclear what the logical unit for an imaging experiments will be. More details under “Unresolved questions”. +1. Still unclear what the logical unit for an imaging experiments will be. More details under "Unresolved questions". 1. Still unclear how to create logical units if data files from the same library preparation are submitted at different times (in more than one submission envelope). Is this a complex update to an existing logical unit? Or a new logical unit that re-uses many metadata files? -Data consumers will benefit from the metadata model now aligning with INSDC “experiments” and “runs”. This alignment will also make archiving to ENA/SRA more straightforward. +Data consumers will benefit from the metadata model now aligning with INSDC experiments and runs. This alignment will also make archiving to ENA/SRA more straightforward. - Library preparation is equivalent to an INSDC experiment - Single set of files from a library preparation is equivalent to an INSDC run | FILE NAME | INPUT LIBRARY PREPARATION ID | INSDC EXPERIMENT ACCESSION | INSDC RUN ACCESSIONS | |:-|:-|:-|:-| -| `sequence_file.file_core. file_name` | `library_preparation.biomaterial_core. biomaterial_id` | `process.insdc_experiment. insdc_experiment_accession` | `sequence_file. insdc_run_accessions` | +| `sequence_file.file_core. file_name` | `library_preparation.biomaterial_ core.biomaterial_id` | `process.insdc_experiment. insdc_experiment_accession` | `sequence_file. insdc_run_accessions` | | SRR7159837_1.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159837 | | SRR7159837_2.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159837 | | SRR7159838_1.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159838 | | SRR7159838_2.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159838 | -**Table 3**: An example set of sequence file metadata based on Fig. 1B experimental design showing how INSDC experiment accession aligns with library preparation ID and how INSDC run accession aligns with individual sets of files (i.e. a “run”). User-friendly field names in row 1; fully-qualified programmatic field names in row 2. **NB**: The `process.insdc_experiment.insdc_experiment_accession` field can be moved to the library preparation entity. +**Table 3**: An example set of sequence file metadata based on Fig. 1B experimental design showing how INSDC experiment accession aligns with library preparation ID and how INSDC run accession aligns with individual sets of files (i.e. a run). User-friendly field names in row 1; fully-qualified programmatic field names in row 2. **NB**: The `process.insdc_experiment.insdc_experiment_accession` field can be moved to the library preparation entity. --- @@ -367,12 +365,12 @@ Data consumers will benefit from the metadata model now aligning with INSDC “e #### Data Processing Pipelines - Subscription queries will reference a library preparation entity instead of cell suspension entity, where appropriate. -- If multiple-bundles:1-workflow remains a use case, pipelines can query for all “bundles” with a particular library preparation UUID in order to trigger a single workflow. +- If multiple-bundles:1-workflow remains a use case, pipelines can query for all bundles with a particular library preparation UUID in order to trigger a single workflow. #### Data Brower - Is anything coded against cell suspension? -- How will this affect displaying rows in “Sample” view? +- How will this affect displaying rows in Sample view? #### Matrix Service @@ -395,12 +393,12 @@ Data consumers will benefit from the metadata model now aligning with INSDC “e #### To solve during RFC process - How will the DCP enforce the requirement of having the library preparation entity in every logical unit (i.e. bundle)? -- How will current production sequencing datasets be updated? All of the current datasets will need to be updated as all of them will need to have library preparation entities added. This is a complex update - change in “bundle” structure by adding a library_preparation_0.json entity - so it can not be done with simple AUDR. If the update happens after GA but before complex AUDR, the UUIDs will have to be maintained somehow. +- How will current production sequencing datasets be updated? All of the current datasets will need to be updated as all of them will need to have library preparation entities added. This is a complex update - change in "bundle" structure by adding a `library_preparation_0.json` entity - so it can not be done with simple AUDR. If the update happens after GA but before complex AUDR, the UUIDs will have to be maintained somehow. #### To solve at future date -- How might per-plate logical units be defined? The plate ID field is currently in the cell suspension schema. Creating a logical unit of “all data per plate” will not be made easier or harder if this proposal is accepted. -- Could this approach work in an analogous way for imaging experiments? I.e. anchor the logical unit on imaged specimen? It sounds like we don’t yet know yet what the “logical” unit is for image-based experiments. One option to anchor on the imaged specimen, but there are also other options. +- How might per-plate logical units be defined? The plate ID field is currently in the cell suspension schema. Creating a logical unit of "all data per plate" will not be made easier or harder if this proposal is accepted. +- Could this approach work in an analogous way for imaging experiments? I.e. anchor the logical unit on imaged specimen? It sounds like we do not yet know yet what the "logical" unit is for image-based experiments. One option to anchor on the imaged specimen, but there are also other options. - What are the naming conventions for user-supplied IDs and system-supplied, globally unique UUIDs? What are the rules for the names of the fields themselves? What are the rules for what values are valid for these fields? Related to metadata-schema issue [#733](https://github.com/HumanCellAtlas/metadata-schema/issues/733). ### Drawbacks and Limitations [optional] @@ -411,9 +409,9 @@ This RFC is based on the current bundle structure and DCP design. If bundle stru ### Prior Art [optional] -Previous [Library preparation entity RFC draft](https://docs.google.com/document/d/15-UkjbAlGDGhOPw1Zu09YdoaGw2Zeq5Y3uDlGsrz6pw/edit) -[Slides](https://drive.google.com/open?id=1vyw6N7qn24qBFAMoKL3nXLHcqpqYFq3Y) from June 2019 DCP F2F meeting outlining library preparation issue from data processing pipelines perspective. -[Slides](https://docs.google.com/presentation/d/1aV7Lrq8SWNxp3Jy98RxUh2SJrLiFPO4Y6AyaH3F1EUI/edit#slide=id.g599322258d_0_8) from June 2019 DCP F2F meeting outlining what is contained in bundles (logical units). +- Previous [Library preparation entity RFC draft](https://docs.google.com/document/d/15-UkjbAlGDGhOPw1Zu09YdoaGw2Zeq5Y3uDlGsrz6pw/edit) +- [Slides](https://drive.google.com/open?id=1vyw6N7qn24qBFAMoKL3nXLHcqpqYFq3Y) from June 2019 DCP F2F meeting outlining library preparation issue from data processing pipelines perspective. +- [Slides](https://docs.google.com/presentation/d/1aV7Lrq8SWNxp3Jy98RxUh2SJrLiFPO4Y6AyaH3F1EUI/edit#slide=id.g599322258d_0_8) from June 2019 DCP F2F meeting outlining what is contained in bundles (logical units). ### Alternatives [optional] From 14b7200b18f854d9cbdac8d2a84f274bddaed375 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 17:24:42 +0100 Subject: [PATCH 21/36] Fixed author formatting. --- rfcs/text/0000-rfc-library-preparation.md | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 764e3811..bc46c93a 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -14,9 +14,9 @@ The current HCA DCP metadata model explicitly represents cell suspensions (singl *Recommended format for Authors:* - `[Mallory Freeberg](mailto:mfreeberg@ebi.ac.uk)` + [Mallory Freeberg](mailto:mfreeberg@ebi.ac.uk) - `[Laura Clark](mailto:laura@ebi.ac.uk)` + [Laura Clark](mailto:laura@ebi.ac.uk) `[Name](mailto:username@example.com)` From 66dac517bbd6c4cbeef85c3b8e3dada5b25cc88b Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 16 Jul 2019 17:26:21 +0100 Subject: [PATCH 22/36] Added graffle image files. --- .../0000-lib_prep_rfc_figs.graffle/data.plist | Bin 0 -> 34922 bytes .../image1.tiff | Bin 0 -> 1052726 bytes .../image2.tiff | Bin 0 -> 10244726 bytes .../image3.tiff | Bin 0 -> 255102 bytes 4 files changed, 0 insertions(+), 0 deletions(-) create mode 100644 rfcs/images/0000-lib_prep_rfc_figs.graffle/data.plist create mode 100644 rfcs/images/0000-lib_prep_rfc_figs.graffle/image1.tiff create mode 100644 rfcs/images/0000-lib_prep_rfc_figs.graffle/image2.tiff create mode 100644 rfcs/images/0000-lib_prep_rfc_figs.graffle/image3.tiff diff --git a/rfcs/images/0000-lib_prep_rfc_figs.graffle/data.plist b/rfcs/images/0000-lib_prep_rfc_figs.graffle/data.plist new file mode 100644 index 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2019 17:31:10 +0100 Subject: [PATCH 23/36] Table formatting fixes. --- rfcs/text/0000-rfc-library-preparation.md | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index bc46c93a..740c86fe 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -276,9 +276,9 @@ Adding a new first-class biomaterial entity has the potential to add a lot of co Data contributors will supply a project-wide unique ID for each library preparation in the Sequence file tab using the `library_preparation.biomaterial_core.biomaterial_id` field in place of the current `sequence_file.library_prep_id` field. When the spreadsheet is imported, the Ingestion Service will generate a globally unique UUID for each library preparation entity. Unlike other biomaterials, there would not be a separate spreadsheet tab for library preparation entities for two reasons. 1. Adding a spreadsheet tab adds considerable complexity for data contributors. 2. There are only two fields that need to be supplied by the data contributor for which generating a whole new tab creates high overhead, especially considering one of the fields replaces a field already in the Sequence file tab. Without the ability to automatically infer `library_preparation.biomaterial_core.ncbi_taxon_id`, this field would need to be added to the Sequence file tab and would need to be filled in by contributors. For example: -| FILE NAME (Required) | *INPUT* CELL SUSPENSION ID (*Required*) | INPUT LIBRARY PREPARATION ID (Required) | *NCBI TAXON ID (Required)* | +| FILE NAME (Required) | INPUT CELL SUSPENSION ID (*Required*) | *INPUT* LIBRARY PREPARATION ID (Required) | *NCBI TAXON ID (Required)* | |:-|:-|:-|:-| -| `sequence_file.file_core. file_name` | `cell_suspension.biomaterial_ core.biomaterial_id` | *`library_preparation.biomaterial_ core.biomaterial_id`* | *`library_preparation.biomaterial_ core.ncbi_taxon_id`* | +| `sequence_file.file_ core.file_name` | `cell_suspension.biomaterial_ core.biomaterial_id` | *`library_preparation.bio material_core.biomaterial_id`* | *`library_preparation.bio material_core.ncbi_taxon_id`* | | SRR7159837_1.fastq.gz | cell_suspension_1 | library_preparation_1 | *9606* | | SRR7159837_2.fastq.gz | cell_suspension_1 | library_preparation_1 | *9606* | | SRR7159838_1.fastq.gz | cell_suspension_1 | library_preparation_2 | *9606* | From 9106e72f1ee44056b807a8f4689fba6cab495555 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Wed, 17 Jul 2019 09:33:48 +0100 Subject: [PATCH 24/36] Added section about removing old field. --- rfcs/text/0000-rfc-library-preparation.md | 13 +++++++++++-- 1 file changed, 11 insertions(+), 2 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 740c86fe..706cb4ac 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -177,7 +177,9 @@ This proposal will require modifications to some DCP components (see "Implementa ### Metadata schema for library preparation entity -**Standard biomaterial fields**. The new `library_preparation.json` schema will follow the schema structure indicated in the Metadata Schema Style Guide and will include the standard properties (fields) for biomaterial type schemas, including: +#### Standard biomaterial fields + +The new `library_preparation.json` schema will follow the schema structure indicated in the Metadata Schema Style Guide and will include the standard properties (fields) for biomaterial type schemas, including: ``` "describedBy": { @@ -268,7 +270,14 @@ The required fields for the library preparation entity will be: - `library_preparation.biomaterial_core.biomaterial_id` - `library_preparation.biomaterial_core.ncbi_taxon_id` -**New library preparation-specific fields**. The new `library_preparation.json` schema will additionally include a field for capturing INSDC experiment accessions if the project is already archived. This field (`process.insdc_experiment.insdc_experiment_accession`) generally represents a single library preparation in the archives (ENA, SRA, DDJB), and currently lives awkwardly in a process module for historical reasons. Additional library preparation-specific fields could be added in the future. +#### Added/changed library preparation-specific fields + +The new `library_preparation.json` schema will include a field for capturing INSDC experiment accessions if the project is already archived. This field (`process.insdc_experiment.insdc_experiment_accession`) generally represents a single library preparation in the archives (ENA, SRA, DDJB). This field currently lives awkwardly in a process module for historical reasons and should be moved to the new library preparation schema. + +The current `sequence_file.library_prep_id` field will be removed. It is no longer needed because the user-supplied `library_preparation.biomaterial_core.biomaterial_id` field captures this information. + +Additional library preparation-specific fields could be added in the future to support data contributors and consumers. + ### Projection of library preparation entity in metadata spreadsheet From 8e05dbabcefc1ea89bd3ce11aff85e5ab7b5c295 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Wed, 17 Jul 2019 09:42:01 +0100 Subject: [PATCH 25/36] Fixed table formatting. --- rfcs/text/0000-rfc-library-preparation.md | 4 ++-- 1 file changed, 2 insertions(+), 2 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 706cb4ac..f4a3adeb 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -287,7 +287,7 @@ Data contributors will supply a project-wide unique ID for each library preparat | FILE NAME (Required) | INPUT CELL SUSPENSION ID (*Required*) | *INPUT* LIBRARY PREPARATION ID (Required) | *NCBI TAXON ID (Required)* | |:-|:-|:-|:-| -| `sequence_file.file_ core.file_name` | `cell_suspension.biomaterial_ core.biomaterial_id` | *`library_preparation.bio material_core.biomaterial_id`* | *`library_preparation.bio material_core.ncbi_taxon_id`* | +| `sequence_file.file_ core.file_name` | `cell_suspension.biomat erial_core.biomaterial_id` | *`library_preparation.biomat erial_core.biomaterial_id`* | *`library_preparation.biomat erial_core.ncbi_taxon_id`* | | SRR7159837_1.fastq.gz | cell_suspension_1 | library_preparation_1 | *9606* | | SRR7159837_2.fastq.gz | cell_suspension_1 | library_preparation_1 | *9606* | | SRR7159838_1.fastq.gz | cell_suspension_1 | library_preparation_2 | *9606* | @@ -332,7 +332,7 @@ Data consumers will benefit from the metadata model now aligning with INSDC expe | FILE NAME | INPUT LIBRARY PREPARATION ID | INSDC EXPERIMENT ACCESSION | INSDC RUN ACCESSIONS | |:-|:-|:-|:-| -| `sequence_file.file_core. file_name` | `library_preparation.biomaterial_ core.biomaterial_id` | `process.insdc_experiment. insdc_experiment_accession` | `sequence_file. insdc_run_accessions` | +| `sequence_file.file_ core.file_name` | `library_preparation.biomat erial_core.biomaterial_id` | `process.insdc_experiment. insdc_experiment_accession` | `sequence_file. insdc_run_accessions` | | SRR7159837_1.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159837 | | SRR7159837_2.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159837 | | SRR7159838_1.fastq.gz | library_preparation_1 | SRX3364233 | SRR7159838 | From 7ffd25b1cffdc5c3b39ee9bb5b6a11ca8a014779 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Wed, 17 Jul 2019 19:01:45 +0100 Subject: [PATCH 26/36] Updated graffle file. --- .../0000-lib_prep_rfc_figs.graffle/data.plist | Bin 34922 -> 34924 bytes 1 file changed, 0 insertions(+), 0 deletions(-) diff --git a/rfcs/images/0000-lib_prep_rfc_figs.graffle/data.plist 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zL^6q_x5oq%sz}5f)C^-Om?Ul%1*Ib7{g{$I5AvqnZeH1q!33R8$f+rj6~;DG Date: Wed, 24 Jul 2019 16:27:36 +0100 Subject: [PATCH 27/36] Updated ES query. --- rfcs/text/0000-rfc-library-preparation.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index f4a3adeb..b8f22b62 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -312,7 +312,7 @@ Data consumers will be able to find all data that need to be processed together "es_query": { "query": { "match": { - "files.library_preparation_0_json.provenance.document_id": "fd420633-6b3d-4361-ac0d-b57b8d9214d7" + "files.library_preparation_json.provenance.document_id": "fd420633-6b3d-4361-ac0d-b57b8d9214d7" } } } From 4fb9d46120593fe3af5e2524a213bdacdbc6675d Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Thu, 25 Jul 2019 13:05:27 +0100 Subject: [PATCH 28/36] Updated definition of LP biomaterial. --- rfcs/text/0000-rfc-library-preparation.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index b8f22b62..fc025a43 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -33,7 +33,7 @@ The current HCA DCP metadata model explicitly represents cell suspensions (singl **Logical unit** - a set of metadata and data files that are consumed together to provide a context for understanding, processing, and interpreting data. In the HCA DCP, a logical unit is instantiated by a primary or secondary “bundle”, although the meaning of bundle is not stable or clearly defined at the moment. -**Library preparation** (biomaterial) - A collection of DNA fragments that have been prepared for sequencing from a single biological sample. DNA fragments can be prepared from genomic DNA or transcribed from RNA and are usually ligated to adapter and barcode oligonucleotides. +**Library preparation** (biomaterial) - A collection of DNA fragments that have been prepared for sequencing. DNA fragments can be prepared from genomic DNA or transcribed from RNA and are usually ligated to adapter and barcode oligonucleotides. **Library preparation protocol** - A protocol followed that converts endogenous RNA or DNA molecules into a form (cDNA) that can be sequenced. Often this protocol involves transcribing RNA to cDNA and ligating adapter and barcode oligonucleotides to the cDNA. From f1cd7fb6676453ad9e11e05b97d99312d7984440 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Mon, 29 Jul 2019 14:00:09 +0100 Subject: [PATCH 29/36] Added caveat about ncbi_taxon_id. --- rfcs/text/0000-rfc-library-preparation.md | 4 +++- 1 file changed, 3 insertions(+), 1 deletion(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index fc025a43..5710103b 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -214,7 +214,7 @@ Where the `biomaterial_core.json` schema contains the following properties: "biomaterial_description": { "description": "A general description of the biomaterial.", }, -"ncbi_taxon_id" : { +"ncbi_taxon_id‡" : { "description": "A taxonomy ID (taxonID) from NCBI.", }, "biosamples_accession": { @@ -235,6 +235,8 @@ Where the `biomaterial_core.json` schema contains the following properties: ``` > *Consider removing `genotype` and `HDBR_accession` from core as they don't make sense here and in other biomaterials. +> ‡The field `ncbi_taxon_id` will remain required as per this RFC. In the future, we can develop tooling that will automatically populate this field from, e.g., the value entered for `ncbi_taxon_id` in the `donor_organism.json` schema. + And the `provenance.json` schema contains the following properties (all supplied by Ingestion Service): ``` From 694201e3f02fbb91c60adf1f88f298dd3d102a40 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 30 Jul 2019 10:46:18 +0100 Subject: [PATCH 30/36] Added Justin as Shepherd. --- rfcs/text/0000-rfc-library-preparation.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 5710103b..6ad75103 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -25,7 +25,7 @@ The current HCA DCP metadata model explicitly represents cell suspensions (singl *Recommended format for Shepherds:* - `[Name](mailto:username@example.com)` + [Justin Clark-Casey](mailto:justincc@ebi.ac.uk) ## Motivation From 96a831747f81c6cd24a2d465af571b2f84198c22 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Tue, 30 Jul 2019 10:47:27 +0100 Subject: [PATCH 31/36] Cleaned up author/shepherd formatting. --- rfcs/text/0000-rfc-library-preparation.md | 7 ------- 1 file changed, 7 deletions(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 6ad75103..a61664bc 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -12,18 +12,11 @@ The current HCA DCP metadata model explicitly represents cell suspensions (singl ## Author(s) -*Recommended format for Authors:* - [Mallory Freeberg](mailto:mfreeberg@ebi.ac.uk) [Laura Clark](mailto:laura@ebi.ac.uk) - `[Name](mailto:username@example.com)` - ## Shepherd -***Leave this blank.** This role is assigned by DCP PM to guide the **Author(s)** through the RFC process.* - -*Recommended format for Shepherds:* [Justin Clark-Casey](mailto:justincc@ebi.ac.uk) From 6e5c9ae9ef58f310b9c46db647c1b235765aec08 Mon Sep 17 00:00:00 2001 From: Mallory Freeberg Date: Wed, 31 Jul 2019 14:02:24 +0100 Subject: [PATCH 32/36] Updated logical unit definition. --- rfcs/text/0000-rfc-library-preparation.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index a61664bc..5c3c972f 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -24,7 +24,7 @@ The current HCA DCP metadata model explicitly represents cell suspensions (singl ### Definitions -**Logical unit** - a set of metadata and data files that are consumed together to provide a context for understanding, processing, and interpreting data. In the HCA DCP, a logical unit is instantiated by a primary or secondary “bundle”, although the meaning of bundle is not stable or clearly defined at the moment. +**Logical unit** - (for the purposes of understanding this RFC; not a suggested new DCP concept) a set of metadata and data files that are consumed together to provide a context for understanding, processing, and interpreting data. In the HCA DCP, a logical unit is instantiated by a primary or secondary "bundle", although the meaning of bundle is not stable or clearly defined at the moment. **Library preparation** (biomaterial) - A collection of DNA fragments that have been prepared for sequencing. DNA fragments can be prepared from genomic DNA or transcribed from RNA and are usually ligated to adapter and barcode oligonucleotides. From 884346ce1d6f557c170dab99f2694a9d201cf679 Mon Sep 17 00:00:00 2001 From: Laura Clarke Date: Mon, 5 Aug 2019 10:31:05 +0100 Subject: [PATCH 33/36] spelling fix in surname --- rfcs/text/0000-rfc-library-preparation.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 5c3c972f..647799d8 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -14,7 +14,7 @@ The current HCA DCP metadata model explicitly represents cell suspensions (singl [Mallory Freeberg](mailto:mfreeberg@ebi.ac.uk) - [Laura Clark](mailto:laura@ebi.ac.uk) + [Laura Clarke](mailto:laura@ebi.ac.uk) ## Shepherd From 2073de7abec85458af7d5f909d315db8c3033651 Mon Sep 17 00:00:00 2001 From: Justin Clark-Casey Date: Thu, 8 Aug 2019 13:39:38 +0100 Subject: [PATCH 34/36] add last call for oversight --- rfcs/text/0000-rfc-library-preparation.md | 2 ++ 1 file changed, 2 insertions(+) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0000-rfc-library-preparation.md index 647799d8..239c840b 100644 --- a/rfcs/text/0000-rfc-library-preparation.md +++ b/rfcs/text/0000-rfc-library-preparation.md @@ -8,6 +8,8 @@ ## Summary +**2019-08-15: Last call for oversight review** + The current HCA DCP metadata model explicitly represents cell suspensions (single cells or multiple cells suspended in some media) but not the sequencing library preparations derived from them. This is creating challenges for contributors, consumers, and DCP implementation teams when submitting, processing, and interpreting sequencing data. Here we propose a solution for explicitly identifying library preparation biomaterials in a sequencing experiment by making them a first-class biomaterial type entity in the metadata standard. ## Author(s) From 41c6957637040887ccab8e1c436f5d4e211798c6 Mon Sep 17 00:00:00 2001 From: Justin Clark-Casey Date: Wed, 21 Aug 2019 17:33:43 +0100 Subject: [PATCH 35/36] Rename 0000-rfc-library-preparation.md to 0010-rfc-library-preparation.md --- ...rfc-library-preparation.md => 0010-rfc-library-preparation.md} | 0 1 file changed, 0 insertions(+), 0 deletions(-) rename rfcs/text/{0000-rfc-library-preparation.md => 0010-rfc-library-preparation.md} (100%) diff --git a/rfcs/text/0000-rfc-library-preparation.md b/rfcs/text/0010-rfc-library-preparation.md similarity index 100% rename from rfcs/text/0000-rfc-library-preparation.md rename to rfcs/text/0010-rfc-library-preparation.md From 68f0c42fc9bd55297be8c241663721271c90b694 Mon Sep 17 00:00:00 2001 From: Justin Clark-Casey Date: Wed, 21 Aug 2019 17:35:27 +0100 Subject: [PATCH 36/36] add link --- rfcs/text/0010-rfc-library-preparation.md | 2 +- 1 file changed, 1 insertion(+), 1 deletion(-) diff --git a/rfcs/text/0010-rfc-library-preparation.md b/rfcs/text/0010-rfc-library-preparation.md index 239c840b..bc70f8b2 100644 --- a/rfcs/text/0010-rfc-library-preparation.md +++ b/rfcs/text/0010-rfc-library-preparation.md @@ -2,7 +2,7 @@ ***Leave this blank until the RFC is approved** then the **Author(s)** must create a link between the assigned RFC number and this pull request in the format:* -`[dcp-community/rfc#](https://github.com/HumanCellAtlas/dcp-community/pull/)` +`[dcp-community/rfc0010](https://github.com/HumanCellAtlas/dcp-community/pull/87)` # Representing sequencing library preparations in the HCA DCP metadata standard

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