Reads trimming

[m4merg]

Hello!

We use PGM for target sequencing. Picture of alignment for single amplicon is shown in attachment. Can someone explain why some reads are trimmed in the 3' end (i.e. reads dont form straight stack in the picture)? I mean at what stage of sequencing it happends. And can be 5' end trimmed? Trimming after sequencing by phred quatility was not done. There is no any strand bias of trimming. Reads are trimmed already in fastq file derived straightly from PGM, so it is not effect of any soft or tool.Trimming length differs from read to read, it is not constant.

Thanks in advance!

[Brian Bushnell]

Were these reads paired?

[m4merg]

Were these reads paired?

Single read sequencing was performed. Reads are not paired

[Brian Bushnell]

So... are you certain that the ones trimmed on the left side are mapped to the plus strand, or might they be mapped to the minus strand? I'm not familiar with that visualizer.

[m4merg]

So... are you certain that the ones trimmed on the left side are mapped to the plus strand, or might they be mapped to the minus strand? I'm not familiar with that visualizer.

Yes, im sure. All reads are trimmed only from 3' end. There are also reverse reads, mapped to the minus strand and trimmed from 3' end, such reads just not shown on picture i've attached. So i've also asked whether 5' end trimming is possible or no.

[Brian Bushnell]

There ARE tools that trim both the 5' and 3' end of reads. What software did you run the data through prior to mapping it?

...also, the mapper could have hard-clipped bases from the read. What mapper did you use, and what command line?

[m4merg]

There ARE tools that trim both the 5' and 3' end of reads. What software did you run the data through prior to mapping it?

...also, the mapper could have hard-clipped bases from the read. What mapper did you use, and what command line?

I know there are tools to trim reads, but as i said I didnt use any of them. Alignment was performed with bwa

bwa index -p $referenceName $referenceName.fasta
bwa mem -t 4 $referenceName $readsFile.fastq > $readsFile.sam 2 > ErrorLog.log

But as i said reads are not trimmed by soft. Reads are trimmed already in fastq file derived straightly from PGM.