SEQanswers

Go Back   SEQanswers > Bioinformatics > Bioinformatics



Similar Threads
Thread Thread Starter Forum Replies Last Post
Genome size estimation with BBTools: what is the right k value? svitlana Bioinformatics 2 07-03-2017 04:27 AM
Error correction in BBTools Sven Bioinformatics 3 06-12-2017 10:54 AM
Additional question regarding BBtools steepale Bioinformatics 3 10-06-2016 12:53 PM
BWA properly paired reads are not really properly paired? metheuse Bioinformatics 0 06-19-2015 11:29 AM

Reply
 
Thread Tools
Old 11-06-2017, 02:53 AM   #1
sghignone
Junior Member
 
Location: Italy

Join Date: Nov 2008
Posts: 6
Default bbtools: how to properly set up rqcfilter

Dear all,
I've recently obtained some PE RNA-seq data from JGI. They have been cleaned using the rqcfilter tool (v. 37.17). Since I would like to uniform the cleaning method extending it to other previously obtained Illumina data, I would like to set up my server to be able to run rqcfilter. I'm aware that this script has been hardcoded to run on on Genepool...but it worth to give a try.

Here follow the steps run by the RQCfilter, as deduced by the reproduce outputs, on my PE data:

>clumpify.sh pigz=t unpigz=t zl=4 reorder in1=OMISSIS.fastq.gz out1=TEMP_CLUMP_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz passes=1

>bbduk.sh ktrim=r ordered minlen=49 minlenfraction=0.33 mink=11 tbo tpe rcomp=f overwrite=true k=23 hdist=1 hdist2=1 ftm=5 pigz=t unpigz=t zl=4 ow=true in1=TEMP_CLUMP_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz out1=TEMP_TRIM_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz rqc=hashmap outduk=ktrim_kmerStats1.txt stats=ktrim_scaffoldStats1.txt loglog ref=adapters2.fa

>rm TEMP_CLUMP_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz

>bbduk.sh maq=10,0 trimq=6 qtrim=r ordered overwrite=true maxns=1 minlen=49 minlenfraction=0.33 k=25 hdist=1 pigz=t unpigz=t zl=6 cf=t barcodefilter=crash ow=true in1=TEMP_TRIM_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz out1=TEMP_FILTER1_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz outm=synth1.fq.gz rqc=hashmap outduk=kmerStats1.txt stats=scaffoldStats1.txt loglog ref=pJET1.2.fasta

>rm TEMP_TRIM_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz

>bbduk.sh ordered overwrite=true k=20 hdist=1 pigz=t unpigz=t zl=6 ow=true in1=TEMP_FILTER1_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz out1=TEMP_FILTER2_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz outm=synth2.fq.gz outduk=kmerStats2.txt stats=scaffoldStats2.txt loglog ref=short.fa

>rm TEMP_FILTER1_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz

>bbmap.sh ordered quickmatch k=13 idtag=t printunmappedcount ow=true qtrim=rl trimq=10 untrim build=1 null path= pigz=t unpigz=t zl=6 minid=.95 idfilter=.95 maxindel=3 minhits=2 bw=12 bwr=0.16 null maxsites2=10 tipsearch=0 in1=TEMP_FILTER2_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz outu1=TEMP_MICROBE_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz outm=microbes.fq.gz scafstats=commonMicrobes.txt

>rm TEMP_FILTER2_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz

>bbmap.sh ordered k=14 idtag=t usemodulo printunmappedcount ow=true qtrim=rl trimq=10 untrim kfilter=25 maxsites=1 tipsearch=0 minratio=.9 maxindel=3 minhits=2 bw=12 bwr=0.16 fast=true maxsites2=10 outm=human.fq.gz path= refstats=refStats.txt pigz=t unpigz=t zl=9 in1=TEMP_MICROBE_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz outu1=OMISSIS.anqrpht.fastq.gz

>rm TEMP_MICROBE_8352bfa6997ae09cc4039681816c_OMISSIS.anqrpht.fastq.gz

>bbmerge.sh loose overwrite=true in1=OMISSIS.anqrpht.fastq.gz ihist=ihist_merge.txt outc=cardinality.txt pigz=t unpigz=t zl=9 adapters=adapters2.fa

>kmercountexact.sh overwrite=true in1=OMISSIS.anqrpht.fastq.gz khist=khist.txt peaks=peaks.txt unpigz=t

I have highlighted in bold the files that, at a first glance, are missing. Where can I find them? while it's pretty clear what could be the content of adapters2.fa and pJET1.2.fasta, what's the content of the short.fa file?

about the contaminant removal, should I retrieve also human and cat/dog/mouse/microbial references or the script does it itself?

Thanks for any hints you may provide!
Cheers,
Stefano
sghignone is offline   Reply With Quote
Old 11-06-2017, 03:56 AM   #2
GenoMax
Senior Member
 
Location: East Coast USA

Join Date: Feb 2008
Posts: 6,550
Default

I think you are better off asking this question to JGI tech support who gave you this data.

@Brian Bushnell (author of BBMap) works at JGI and contributes regularly on this forum. He has been busy with something for last few days and I have not seen him here. You could also try emailing him directly.
GenoMax is offline   Reply With Quote
Old 11-06-2017, 04:24 AM   #3
sghignone
Junior Member
 
Location: Italy

Join Date: Nov 2008
Posts: 6
Default

I supposed he has been busy...I 've wrote him last week and got no replies, that's why I posted here. I'll give another try by email and with JGI staff.
Thanks!
s.-
sghignone is offline   Reply With Quote
Reply

Tags
bbduk, bbmap, bbtools, clumpify, rqcfilter

Thread Tools

Posting Rules
You may not post new threads
You may not post replies
You may not post attachments
You may not edit your posts

BB code is On
Smilies are On
[IMG] code is On
HTML code is Off




All times are GMT -8. The time now is 11:22 AM.


Powered by vBulletin® Version 3.8.9
Copyright ©2000 - 2017, vBulletin Solutions, Inc.
Single Sign On provided by vBSSO