Separate Broken Pairs

Question

[Tom]

I am currently preprocessing my paired-end reads and was wondering if there is an efficient way to remove broken pairs from my sequence file.

Thanks for the help.

[Obi Griffith]

Some more details would be good. What format is your RNA-seq data in and how has it been processed so far? And, how would you like to define "broken pairs"?

Answer 1

[Istvan Albert]

This post might help: Filtering paired end reads

Answer 2

[Tom]

It is in fastq format and following QC filtering with fastx I end up single reads that no longer have a pair. It seems that these single reads are messing up my assemblies in velvet. Recently, I tried using sickle to filter and it seems to have resolved the issue.

Thanks for the help.

Answer 3

[Ian]

Picard tools has a handy webpage for working out the meaning of SAM flags (and combinations of flags):

http://picard.sourceforge.net/explain-flags.html

These can be used with 'samtools view -f N', where N is the flag you want to filter your reads with.

E.g. to extract all the properly paired reads:

samtool view -S -f 2 -o proper-paired-reads.sam input.sam

Answer 4

[Tom]

That is a great tool!

Cheers