Does BBDuk work on concatenated fastq files

[Pluto]

Basically, I have around 200 samples and wanted to sequence at a depth of 50 million reads per sample. This was not possible on one run so I chose to do all 200, 14 times. I have concatenated all the R1 together and all the R2 together from one sample and so on.... so now I have 200 R1 and 200 R2.

I was wondering if BBDuk can deal with these files as each file is made up of 14 fastq output with 14 headings.

Thank you in advance.

[GenoMax]

BBduk will work fine. Hopefully you concatenated the files in exactly the same order for both R1/R2 files.

[Pluto]

Adaptor trimming is not working

Thank you for getting back to me. The adaptor trimming is not working sadly.

This is what my script looks like:

Ordered=t #Set to true to output reads in same order as input
Ktrim=r #once a reference kmer is matched in a read, that kmer and all the bases to the right will be trimmed
K=21 #specifies the kmer size
Mink=8 #"mink" allows it to use shorter kmers at the ends of the read
Hdist=2 #number of permitted mismatches


for Prefix in `ls -1 *_R1.fastq.gz | sed 's/_R1.fastq.gz//'`
do

bbduk.sh -Xmx128g in1=$Prefix\_R1.fastq.gz in2=$Prefix\_R2.fastq.gz out1=$Prefix\_clean_R1.fastq.gz out2=$Prefix\_clean_R2.fastq.gz ref=$adapters ordered=$Ordered ktrim=$Ktrim k=$K mink=$Mink hdist=$Hdist tpe tbo

done

Remember my R1 and R2 files consist of concatenated sequences from different runs. Do you think this could be the reason?

Many thanks

[GenoMax]

As I said before as long as the files are concatenated in same order AND they had the same number of reads in sync across R1/R2 files to begin with this should work without any problems. If things are not working you need to make sure that the reads in your files are in sync. You can check on that using a different bbtool called "repair.sh".

BBduk.sh needs very little memory there is no need to assign 128G for this job. 4G would be perfectly fine.