Question: RNA-seq stranded protocol
0
gravatar for rubic
6.5 years ago by
rubic200
United States
rubic200 wrote:

Hi,

 
I have RNA-seq data that was generated with Illumina TruSeq stranded protocol, according to which, as far as I understand, first mate reads are supposed to map to the same strand of the transcript and second mate reads are supposed to map to the opposite strand of the transcript. Assuming this is correct and eliminating any read-pair mappings that do not obey this condition I get very few read mappings. I noticed that I have a lot of read-pair mappings where the opposite to my condition is true. I.e., the left mate maps to the opposite strand of the transcript and the right mate maps to the same strand of the transcript. Are these read-pair mappings reliable? Should I not discard them?
rna-seq • 1.8k views
ADD COMMENTlink modified 6.5 years ago by Devon Ryan98k • written 6.5 years ago by rubic200

cross-posted: http://seqanswers.com/forums/showthread.php?t=44754

ADD REPLYlink written 6.5 years ago by Pierre Lindenbaum133k
2
gravatar for Devon Ryan
6.5 years ago by
Devon Ryan98k
Freiburg, Germany
Devon Ryan98k wrote:

I recall that the Illumina TruSeq kit is a dUTP-based protocol, so the strand would be that of read #2.

ADD COMMENTlink modified 6.5 years ago • written 6.5 years ago by Devon Ryan98k

So according to your answer all valid read-pair mappings should obey the rule that the first mate maps to the opposite strand and the the second mate maps to the original strand, and the any other read-pair mappings (such as the opposite pattern) should be discarded?

ADD REPLYlink written 6.5 years ago by rubic200

Correct   

ADD REPLYlink written 6.5 years ago by Devon Ryan98k
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