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Old 04-16-2020, 02:28 AM   #1
maxz411
Junior Member
 
Location: Rochester, NY

Join Date: Feb 2020
Posts: 7
Default Incorrect bbmap quality trimming

I am using bbmap to trim bases below a certain quality threshold in a FASTQ file. The file looks fine to me in terms of quality scores, but when I run bbmap it trims almost all the bases away and seems to leave just one base per read, despite most of the bases meeting the quality threshold.

I am not sure if this makes any difference, but this is a FASTQ file generated by merging overlapping paired-end reads and adjusting quality scores to reflect this with SeqPrep. As a result, most of the quality scores are high (lowercase ASCII characters) and I am setting a trim threshold of 59. Nonetheless, many more of the bases in the file should meet this threshold than what bbmap is outputting.

In case it is helpful, I pasted my command below. I also pasted a few lines of the FASTQ input.

Code:
./reformat.sh in=./samples/B10C_merge.fq out=./samples/B10C_trim.fq qtrim=rl trimq=59 qin=33 qout=33
Code:
@M02713:293:GW200329AmpliconEZC0318:1:2105:15470:1423 1:N:0:TGATCACG+AGGCGAAG
GATCTGTTGCTGCCCCAGGATGTTGAGGAGTTTTTTGAAGGCCCAAGTGAAGCCCTCCAAGTGTTAGGAGCTCCTGCAGCACAGGCCCTGCCCCAGCTACTCCATGGCCCCCGTCATCTTTTGTCCCTTCTCAAAAAACTTACCAGGGCAACTATGGCTTCCACCTGGGCTTCCTGCAGTC
+
hhiijlllllllmmnnnnnnnnooooooomnooonnnmnoooonnoooooonnnooonoooooooooooonlooooooooonnnnmmmnnnnonnnoooooooonnnnnnlmnnnnnoononnonoononnnnnooolnooooooonnnnnoooooooonnnnnnnnnmmmjmmmmijjij
@M02713:293:GW200329AmpliconEZC0318:1:2105:18370:1751 1:N:0:TGATCACG+AGGCGAAG
TTTGATGATGGCTGTCATGTCTGGGAGCCTGTGGCTGAAGAAAAAGGAGGAGAGAGATGGCAGAAGCTGCTGGTGGCGGGGCTTCTTCTGCAGGATGGAAATGGCTCTGGACTTGGCGGTGGCTGATGCCCCTCGCTCTGCTGCCGCTTGGTTCTGGACAGCAGCCGGGTAATGGCTGCTGCGGCGGCTGCTGGATGGTTGCAGCGACTGGGCCTGCTTCTCCTCAGCAGCCA
+
fggggkkkkkkkllllllkgllmmmmlllmmmgmmmlmmmmmmmmlllmlmmnmmmmnnnmnmmnmnnnmlmnnnmnmmnmnmnoonooomooomoonnmonooojnnolnonnnnlnmbhkojmWWljnnmkmjiUjcmlllnniigllnahWaWmWmmgkhllmmnn`jninnmikehhkagdTj`T`UUa`T`WbWaTThhh`TgfgklekkkeVhVV[gU^UQZPZQfO
@M02713:293:GW200329AmpliconEZC0318:1:2105:12451:2026 1:N:0:TGATCACG+AGGCGAAG
TTTGATGATGGCTGTCATGGCAAAGGGAGGAGGACAGGCTTCTCCGTCCCCAGGAAGCAACTGGAGGCCCAGCTGAGCCCAGCTCTGCCTCAGCTTCCCCATCTGTAAAATGGGGTGATGGGCACCAGGCGGTAGGTGCAGCCTCACTGTCTTCTTGCCCCCAGCGGAGCTGATGGAGCGGGCCGCGGTGCCACCCCTTTGGCCGGCCCTGTACCCACCAGGCCGCAGCTCCCTGCACCACGCCCAGCAGCTGCAGCTCTTCTCCTCAGCAGCC
+
CABCCFFFFFFFGGGGGGGGGGHHmmlmllllllmmhllmmmmmkammllmmllmmlmmmmmlmmmljlllmmlmmmmllmmmmmmmmmmmmmmmmmlmmmmmmmmmmmmmmmmlllklmnnnmmmnngnmmmkmmnnnknnooooomonoonljnnnnnlmmmnnnnnnnoonlmknnmlmmmmmlmmdhjTllmmmmllUhllgkmlmmmlllmlmllllllllmllllmmmmklllllmlkmklmHGHHGGGGGGGGGGFCFFFFCCCCCB
@M02713:293:GW200329AmpliconEZC0318:1:2105:19453:2143 1:N:0:TGATCACG+AGGCGAAG
GAGGGGCATCAGCCACCGCCAAGTCCAGAGCCATTTCCATCCTGCAGAAGAAGCCCCGCCACCAGCAGCTTCTGCCATCTCTCTCCTCCTTTTTCTTCAGCCACAGGCTCCCAGACATGACAGCCATCATCAA
+
[Ofd[Q]RS^RRib;eimnlnmnjnmnjYiiimmmYmnjVnmi[iomnomoldnnnmnlmimooooomoonnoojomlkjnlnnnonnlkomkjoonnonomomonnlmonmnjnmnnnnjmljmmmmiiiii
@M02713:293:GW200329AmpliconEZC0318:1:2105:11660:2539 1:N:0:TGATCACG+AGGCGAAG
AGCCCCGCCACCAGCAGCTTCTGCCATCTCTCTCCTCCTTTTTCTTCAGCCACAGGCTCCCAGACATGACAGCCATCATCAAA
+
Qbiiijkjkijmjmnnnnnmmnnooonnnnonnnoonlooommoonnooooooonnmoonnnnmmnnnnnnmmmmmmmiifgi
@M02713:293:GW200329AmpliconEZC0318:1:2105:18713:2826 1:N:0:TGATCACG+AGGCGAAG
CTGTTATTGCTAGCGTTTTAGCACAGGTGCAGCTGGTGGAGTCTGGGGGAGGATTGGTCCAGGCTGGAGGCTCTCTGAGACTCTCCTGTGCAGGCTCTTCACCCGCCTTCACTAAACTCGCCGTGGGGTGGTTCCGCCAGGCTCCAGGAAAGGAGCGTGAGTTTGTCGCAGCTTGTGGTTGGAGTGGAAGTGATACATACTATGCGGACTCCGTGAAGGGCCGATCCAGCATCTTCAGAGACAACGCCAAGAACACGGTGTATCTGCAAATGAACAGCCTGAAACCTGAGGACACGGCCATTTATTACTGTGCAGTGAGAGTATGGTGGGCGGGCGATTGGGATACAGAAACGCAGTATGATTACTGGGGCCAGGGGACCCAGGTCACCGTCTCCTCAGAATTCGGTAAGCCTATCCCTAAC
+
BBBBBFFFFFFFFGGEFGGGGGHHHHHGHFHHFHHHGFGGHFHHGHHGGGGGGHHHEFGFHHHHHHGHHGGHHHHHHHHHHHHEHHHGHHGFHGGHHHEHHHHGGFEFGGFFHHHGHHEGGGG@DGFG<@CGGGHGGGGFGGECEHFHEHHAGGHHGDFGGCGFGFFGHGGGGlkjflgUTUghU_lgkl`^kllkgh^lkigejlgjaejlkjlgilhdcjkilahmlllmmlmkmmmkjllkhkKgmHHGGHFGFFHGHHEGDDGFHHGHHHHHFGHHHHFGEEGFFGHFDBGHGGFFHHFHHHFHHGHFHFFHHEHFGFFHGGGEECEEEAFCF3HFGEGFHGHHECFEHFHFHFGD5HHHDFE1GGHFHHHHFFCHFFG3FFAEEFAGHFGHEDBFEGFEBGGGGGFFFFFFFAA>33
@M02713:293:GW200329AmpliconEZC0318:1:2105:11972:2903 1:N:0:TGATCACG+AGGCGAAG
GGGGCTTCTTCTGCAGGATGGAAATGGCTCTGGACTTGGCGGTGGCTGATGCCCCTCGCTCTGCTGCCGCTTG
+
jkikkjjmmmmmnnnnnnnnnnoooooooonoooomnnnnnnnnmoononnmnnmmmmmmnmjjjjklkkkjj
@M02713:293:GW200329AmpliconEZC0318:1:2105:13695:2965 1:N:0:TGATCACG+AGGCGAAG
TGGCTGCTGAGGAGAAGCAGGCCCAGTCGCTGCAACCATCCAGCAGCCGCCGCAGCAGCCATTACCCGGCTGCTGTCCAGAACCAAGCGGCAGCAGAGCGAGGGGCATCAGCCACCGCCAAGTCCAGAGCCATTTCCATCCTGCAGAAGAAGCCCCGCCACCAGCAGCTTCTGCCATCTCTCTCCTCCTTTTTCTTCAGCCACAGGCTCCCAGACATGACAGCCATCATCAAA
+
OZQPQPQTS8_\VV_VT`T`Sh__;TaTVV`TTh<`Ua;aWW`TTTaTS`SV8VTgVimmmka<mkfiUiU_TW`WiWWaaWhUaTTfiVVbbVUjjTgbTmmlWWlWmcWnhcnnnnnnmmkooommmoooonimmYVonmmmnioooollnnnmnmnmnnnmnnnnnmmnnnnnmmnnkmmlmmllmmmmmmoolommmmlmllllmmmllllllllllkkkkkkkhihgg
@M02713:293:GW200329AmpliconEZC0318:1:2105:15588:3299 1:N:0:TGATCACG+AGGCGAAG
TGGCTGCTGAGGAGAAGCAGGCCCAGTCGCTGCAACCATCCAGCAGCCGCCGCAGCAGCCATTACCCGGCTGCTGTCCAGAACCAAGCGGCAGCAGAGCGAGGGGCATCAGCCACCGCCAAGTCCAGAGCCATTTCCATCCTGCAGAAGAAGCCCCGCCACCAGTAGCTTCTGCCATCTCTCTCCTCCTTTTTCTTCAGCCACAGGCTCCCAGACATGACAGCCATCATCAAA
+
4ZOPP\6TT8^\`9UUU9T_ScaS;TTT`UUUTa<b;U0aTTTT<TbT_`SUUVT`VinmmiaUh`TiUTUiTjaWWajjjWbUbTTSiVnmmljiUTTbTllmWjbWUWVlgcnnlmhnnnnoonmilooooonnonmonommonooonfnnnnmnmnnnnmn`mmnnmmnnnnnmmmmnmmmnmlllmmlmmlmlmmmmmmmlllmmmmllllllllllkkkkkkkhhhhh
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