Hello,
I’ve assembled my v4 reads (MiSeq - 2x300bp) using make.contigs with the following oligos file
primer GTGBCAGCMGCCGCGGTAA GGACTACHVGGGTWTCTAAT v4
My reads have had the index and barcodes removed - but still include primer sequence
I run the command including pdiffs=2, trimoverlap=T
Everything runs fine but I have a question regarding the scrap output (here are a few suspicious ones):
M04603_48_000000000-ATNCR_1_1101_10759_1985 | f fbdiffs=0(match), rbdiffs=0(match) fpdiffs=1(match), rpdiffs=2(match)
TACGTAGGTGGCAAGCGTTGTCCGGAATTATTGGTCGTAAANCGCGNNCAGNCNGATCNGTCAGTCTGTCTTAAAAGTTCGGTGCTTAACCCCGTGATNGGATGGAAANTNCCAATCTAGAGTATCGNAGAGGAAAGTGGAATTCCTAGTGTAGCGGTGAAATNNGTAGATATTAGGAAGAACACCAGTGGCGAAGGCGACTTTCTGGACGAAAACTGACGCTGAGGNGCGAAAGNCAGGGGAGCGAACGGG
M04603_48_000000000-ATNCR_1_1101_23439_1994 | f fbdiffs=0(match), rbdiffs=0(match) fpdiffs=2(match), rpdiffs=1(match)
CGATTAACCCAAGTCAATAGAAGCCGGCGTAAAGAGTGTTTTAGATCACACCCCTCCCNAATAAANCTAAAACTCANCTNAGTTGTAAAAAACTCCAGTTNACACAAAATAGACTACGAAAGTGNCTTTAACATATCTGAACACACAATAGCTAAGACCCAAACTGGG
M04603_48_000000000-ATNCR_1_1101_10238_2844 | f fbdiffs=0(match), rbdiffs=0(match) fpdiffs=2(match), rpdiffs=2(match)
GAGAACAATCTGCTCCAAAATAGAAATAGTGCCCAGAGTGAGAAACCCTGCTCTACAAACCCTACAGTGTACAGGCCAGCCGCCACATCAGAGAATAATCTGCNCCAAACTAGAAATAGTGCCCAGAGTGAGAAACCCTG
fpdiffs and rpdiffs all appear to pass the pdiffs cut off - yet they still don’t make it into the “trim.contigs” file. Also, why do I get an “f” code when my forward primer is a perfect match? Is there another default filter parameter I’m missing?
Thanks