Sequence Trimming without Knowing Primer Sequences

KyuzoSeq · 11-18-2016, 12:59 AM

Hi,

I have an IonTorrent output but the provider doesn't want to give information about primer sequences. Instead it gives a bed file includes regions of interest. So if I do alignment and bedtools intersect than trimming sequences by outside parts which is not in the region, I'm not comfortable this is a good approach. We want to use the output as a data for finding SNP frequencies and for the best approach primer sequences must be trimmed.

So, is there a way to do this job properly? I mean is there a way to trim sequences with no knowledge about primers?

Brian Bushnell · 11-18-2016, 09:54 AM

What kind of experiment are you doing? Is it some kind of exon-capture, or an amplicon approach? Or, why do you expect to see primers in the sequence?

KyuzoSeq · 11-19-2016, 01:30 PM

Quote:

Originally Posted by Brian Bushnell

What kind of experiment are you doing? Is it some kind of exon-capture, or an amplicon approach? Or, why do you expect to see primers in the sequence?

It's basically a variant analysis process. What I need is trimming sequences by primer sequences but we have no information a about primer sequences.

Markiyan · 02-21-2017, 03:33 AM

If you would like to have some idea about the primers you can try:
Converting to fastq format and having a look at fastqc at high kmer settings (-k8 or -k10).

Up the memory to java wm (-Xmx) if you run out of memory for the fastqc application.

Also try assembling a susbset (1k - 10k) of reads matching the primer-like kmers, and having a look at the de novo assembly results in consed or similar program.
Newbler assembler has primer detection capability and may give you some idea about the primers present (check it's log).

Local setup of WWWblast / STARblast or similar tool comes very handy for doing investigations.

grep -c primer *.fasta against the sequences in fasta format may give you an idea about the abundance of the given kmer.

Other options to investigate includes trimmomatic and similar tools.

Also look for high quality unaligned region a the the ends of your reads when examining the mapping results.

Similar Threads
Thread	Thread Starter	Forum	Replies	Last Post
Base modification - Knowing the modified bases fall on coding or non-coding sequences	garethboy	Bioinformatics	0	08-28-2015 12:51 PM
Adapter/primer trimming from RNAseq reads	everestial	General	44	03-05-2015 12:08 PM
Primer sequences emPCR and sequencing primer Ion Torrent	laurakrol	Sample Prep / Library Generation	1	04-25-2014 05:08 AM
Any idea on primer trimming tool?	arkilis	Bioinformatics	5	09-23-2013 10:54 PM
Create one sequence based on overlapping primer sequences in amplicon	ketan_bnf	Bioinformatics	2	09-15-2011 01:33 AM

11-18-2016, 12:59 AM	#1
KyuzoSeq Junior Member Location: Turkey Join Date: Mar 2015 Posts: 9	Sequence Trimming without Knowing Primer Sequences Hi, I have an IonTorrent output but the provider doesn't want to give information about primer sequences. Instead it gives a bed file includes regions of interest. So if I do alignment and bedtools intersect than trimming sequences by outside parts which is not in the region, I'm not comfortable this is a good approach. We want to use the output as a data for finding SNP frequencies and for the best approach primer sequences must be trimmed. So, is there a way to do this job properly? I mean is there a way to trim sequences with no knowledge about primers?

02-21-2017, 03:33 AM	#4
Markiyan Senior Member Location: Cambridge Join Date: Sep 2010 Posts: 116	Use fastq contaminant detection and removal approaches. Do a bit of de novo... If you would like to have some idea about the primers you can try: Converting to fastq format and having a look at fastqc at high kmer settings (-k8 or -k10). Up the memory to java wm (-Xmx) if you run out of memory for the fastqc application. Also try assembling a susbset (1k - 10k) of reads matching the primer-like kmers, and having a look at the de novo assembly results in consed or similar program. Newbler assembler has primer detection capability and may give you some idea about the primers present (check it's log). Local setup of WWWblast / STARblast or similar tool comes very handy for doing investigations. grep -c primer *.fasta against the sequences in fasta format may give you an idea about the abundance of the given kmer. Other options to investigate includes trimmomatic and similar tools. Also look for high quality unaligned region a the the ends of your reads when examining the mapping results.

11-18-2016, 09:54 AM	#2
Brian Bushnell Super Moderator Location: Walnut Creek, CA Join Date: Jan 2014 Posts: 2,707	What kind of experiment are you doing? Is it some kind of exon-capture, or an amplicon approach? Or, why do you expect to see primers in the sequence?