Question: Reads mapped to another chromosome in paired-end data of RNA-seq
0
gravatar for int11ap1
6.9 years ago by
int11ap1420
Barcelona
int11ap1420 wrote:

Hello guys,

I am doing a project in RNA-seq data with some datasets that are paired-end data. After alignment with TopHat2, I have around 1-5% of reads whose mate is mapped to another chromosome. In order to call Cufflinks for annotation of transcripts, would you recommend me to remove those reads mapped to another chromosome (or call again TopHat2 with the option --no-discordant)? Is it a key step?

Thanks in advanced.

rna-seq • 2.9k views
ADD COMMENTlink modified 6.7 years ago by Biostar ♦♦ 20 • written 6.9 years ago by int11ap1420

Sorry this isn't answering your question but instead asking a new one: What tool (or commands) did you use to find out how many reads had mates that mapped to a different chromosome?

ADD REPLYlink modified 6.9 years ago • written 6.9 years ago by Ann2.3k

samtools flagstat FILE.bam

ADD REPLYlink modified 14 months ago by Ram32k • written 6.9 years ago by int11ap1420

I thought bowtie rejected alignments of reads whose mates are too far from one another (than the estimated fragment size). Maybe tophat retains them for splicing event detection?

ADD REPLYlink modified 14 months ago by Ram32k • written 6.9 years ago by ndejay30

I don't think TopHat2 will use read pairs aligning on different chromosomes for annotating transcripts. This is my personnel guess but I am quiet positive about it. So you need not to remove these reads. Still if you want you can use this command:

  1. samtools view -H Your.bam > Your.sam
  2. samtools view Your.bam | awk '$7 == "=" {print}' >> Your.sam
  3. samtools view -bS Your.sam > Your_new.bam
ADD REPLYlink modified 14 months ago by Ram32k • written 6.7 years ago by Ashutosh Pandey12k
Please log in to add an answer.

Help
Access

Use of this site constitutes acceptance of our User Agreement and Privacy Policy.
Powered by Biostar version 2.3.0
Traffic: 1071 users visited in the last hour
_