[khmer] orientation of SE reads

C. Titus Brown ctb at msu.edu
Tue Oct 1 19:48:55 PDT 2013

On Wed, Oct 02, 2013 at 09:12:01AM +0800, bioques wrote:
> Hi Professor Brown,
> I tried to follow the protocol you kindly offer here (https://github.com/ged-lab/khmer-protocols/tree/master/mrnaseq). It really a great help to newbie like me for RNA-Seq analysis. But I got one question about the step after trimming the adapter with Trimmomatic (document in 1-quality.txt). You suggest to combine the SE reads s1_se and s2_se together. Would this hurt the orientation of the reads when doing the assembly (I am going to use Trinity to do the assembly)? I wonder if I should convert the s2_se reads to its reverse complement before combine the two files together?
> Hope to hear from you and thanks in advance!

Hi Nate,

no need to RC the s2_se reads, unless you are dealing with single-strand
mRNAseq (in which case I'm not sure this protocol works for other reasons).
Basic dsDNA sequencing reads do not have any particular orientation unless
you're doing something tricky with mate pairs or single-strand mRNAseq.

C. Titus Brown, ctb at msu.edu

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