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Aligning

Updated: May 11, 2017

Before any alignment, a graph needs to be generated with vargas define. To use default parameters and produce a SAM file,

vargas align -g <graph_def> -U <reads> -S <aligns_out.sam>

where <reads> can be a FASTA/Q or SAM file.

Scoring options

  • --ete Use end to end alignment. This is faster than full local.
  • --ma <INT> Match bonus.
  • --mp <INT> Mismatch penalty (provide a positive number).
  • --rdg <INT,INT> Read gap penalty <Open,Extend>.
  • --rfg <INT,INT> Reference gap penalty <Open,Extend>.
  • --np <INT> Penalty for non-A/C/G/T bases.

Alignment targets

By default, reads will align to the base graph. If the input is in SAM format, specific read groups can be aligned to specific graphs with the -a option. The argument can either (a) be a graph to align to, or (b) A list of target mappings with the format:

vargas align -g <graph_def> -r <reads> -t <out.sam> -a "RG:<tag>:<val>,<subgraph>; ..."

where <tag> can be ID or some auxiliary tag in the SAM Read Group line. RG's with <val> in the <tag> field will be aligned to <subgraph>. Reads that are not associated with a read group are assigned to VAUGRP.

Using a SAM input where an alignment is already defined will enable the reporting of the cf and ts flags.

Assess

If a SAM read file is provided, -s can attempt to match a previous scoring function. Currently Bowtie2, HISAT2, and BWA MEM are supported.

Output

Alignments are written as SAM files. Note CIGAR alignments are not provided. Fields relevant to Vargas are listed below.

  • POS Maximum scoring alignment, only known for end to end alignments.
  • RNAME Maximum scoring sequence.
  • FLAG Reverse complement flag bit if aligned to opposite strand.
  • AS Maximum score.
  • mp Position of the maximum scoring cell.
  • ss Second best score.
  • st Strand of second best score.
  • sp Position of the second highest scoring cell.
  • su Sequence name of the second best score.
  • mc Number of max-score occurrences.
  • sc Number of second-best score occurrences.
  • gd Read group tag. Subgraph aligned to.

vargas convert can be used to extract these fields into a CSV file.

Coordinates

When a graph diverges, parallel nodes can have different lengths. To project the position onto the reference, nodes are anchored to the end of the node. For example:

    012 1234 5678
        ||||
        ACAC
       /    \
    AAC--GT--AAAA
         ||
    012  34  5678

Note how C and T, the ends of the nodes, have the same position.