Enhancement - primer cocktails for a marker

[peterjc]

Some protocols don't just use IUPAC codes to define their forward/reverse amplicon primers, but a cocktail. e.g.

• Arulandhu et al. (2017), our endangered_species worked example) has multiple right primers for COI
• Foster et al. (2022) Oomycete rps10 locus has both multiple left and multiple right primers, see
  http://oomycetedb.cgrb.oregonstate.edu/protocols/metagenomics_primer_protocol.html

Support would need modifying our command line API (e.g. allow --left and --right arguments to be comma or semi-colon separated lists), modifying the SQLite database schema to hold the additional primers, and working out how to pass this to cutadapt while multiplexing.

[peterjc]

See also https://github.com/grunwaldlab/OomyceteDB/blob/master/protocols/tables/metabarcoding_primer_mixing.tsv

rps10 example given as two forward primers:

• GTTGGTTAGAGYAAAAGACT (note Y, meaning C or T - with double the volume)
• GTTGGTTAGAGTAGAAGACT (note T instead of Y, and an A to G change later)

i.e. maps to three unambiguous primers (at same concentration):

• GTTGGTTAGAGCAAAAGACT
• GTTGGTTAGAGTAAAAGACT
• GTTGGTTAGAGTAGAAGACT

Could approximate with the following, but wrongly includes a fourth sequence:

• GTTGGTTAGAGYARAAGACT

Reverse given as seven unambiguous reverse primers (at same concentration):

• ATGCTTAGAAAGATTTGAACT
• ATACTTAGAAAGATTTGAACT
• ATGCTTAGAAAGACTTGAACT
• ATGCTTAGAAAGACTCGAACT
• ATGCCTAGAAAGACTCGAACT
• ATGTTTAGAAAGATTCGAACT
• ATGCTTAGAAAGATTCGAACT

Could approximate with the following, but this is overly diverse:

• ATRYYTAGAAAGAYTYGAACT

[peterjc]

According to https://github.com/grunwaldlab/rps10_barcode/blob/main/raw_data/primer_data.csv the authors did actually use GTTGGTTAGAGYARAAGACT and ATRYYTAGAAAGAYTYGAACT in their own analysis for Foster et al. (2022).