Question: Trimmomatic adapter clipping removes many reads even if "keepBothReads" parameter is selected
0
gravatar for al-ash
3 months ago by
al-ash50
European Union
al-ash50 wrote:

I'm using Trimmomatic/0.33 to proces my raw Illumina reads. I've seen some adapter contamination with FASTQC so I first run Trimmomatic as follows:

java -jar trimmomatic-0.33.jar PE R1.fastq.gz R2.fastq.gz PR1.fastq.gz UPR1.fastq.gz PR2.fastq.gz UPR2.fastq.gz ILLUMINACLIP:TruSeq3-PE-2.fa:2:30:10:1:keepBothReads

TruSeq3-PE-2.fa contains:

>PrefixPE/1
TACACTCTTTCCCTACACGACGCTCTTCCGATCT
>PrefixPE/2
GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCT
>PE1
TACACTCTTTCCCTACACGACGCTCTTCCGATCT
>PE1_rc
AGATCGGAAGAGCGTCGTGTAGGGAAAGAGTGTA
>PE2
GTGACTGGAGTTCAGACGTGTGCTCTTCCGATCT
>PE2_rc
AGATCGGAAGAGCACACGTCTGAACTCCAGTCAC

Trimmomatic reports back:

Input Read Pairs: 39618168 
Both Surviving: 28128581 (71.00%) 
Forward Only Surviving: 8927582 (22.53%) 
Reverse Only Surviving: 47164 (0.12%) 
Dropped: 2514841 (6.35%)

I was surprised that despite selecting the option to keep both reads even if they are palindromic via "keepBothReads" there is only 71% of "Both Surviving" : what might be the reads which did not survive? I can think of them only as entirely consisting of adapter sequences - is that correct? Thanks for any suggestion!

ADD COMMENTlink modified 10 weeks ago by finswimmer5.3k • written 3 months ago by al-ash50
1
gravatar for annsun89
10 weeks ago by
annsun8910
annsun8910 wrote:

Hi, I read the manual and <keepbothreads> is the name of the parameter but it's not how to set it. Instead, try

java -jar trimmomatic-0.33.jar PE R1.fastq.gz R2.fastq.gz PR1.fastq.gz UPR1.fastq.gz PR2.fastq.gz UPR2.fastq.gz ILLUMINACLIP:TruSeq3-PE-2.fa:2:30:10:1:TRUE
ADD COMMENTlink written 10 weeks ago by annsun8910
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