Question: Paired-end RNA sequence - which read is complimentary?
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gravatar for yanweng
3.1 years ago by
yanweng30
yanweng30 wrote:

This may be a basic question. I am doing Illumina stranded, paired-end RNA-seq, and get read1 and read2. Is read1 has the same sequence as the mRNA, or read2 has the same sequence as the mRNA?

rna-seq sequence next-gen gene • 1.2k views
ADD COMMENTlink modified 3.1 years ago • written 3.1 years ago by yanweng30
2
gravatar for Antonio R. Franco
3.1 years ago by
Spain. Universidad de Córdoba
Antonio R. Franco4.0k wrote:

Devon was trying to tell you that if the cDNA you obtain from the RNA is stranded (using the dUTP technique) the read #2 is normally the same that the RNA. That is not the case if you made a stranded cDNA using Illumina TruSeq method (thus read # 1 is the RNA) or if your cDNA is an unstranded. In the latter case you cannot know the origen of your strands

ADD COMMENTlink written 3.1 years ago by Antonio R. Franco4.0k
1
gravatar for Devon Ryan
3.1 years ago by
Devon Ryan88k
Freiburg, Germany
Devon Ryan88k wrote:

The strand that read #2 aligns to is normally the strand from which the fragment arose. That's the case for libraries produced these days (they're all dUTP based) but wasn't always the case.

ADD COMMENTlink written 3.1 years ago by Devon Ryan88k

Is it because we are sequencing cDNA, and cDNA is complementary to mRNA, so R1 is strand-specific to DNA which is complementary to mRNA?

ADD REPLYlink written 3.1 years ago by apelin20470
0
gravatar for yanweng
3.1 years ago by
yanweng30
yanweng30 wrote:

Thanks for your answers! I am using the dUTP technique, so read 2 is the same to the gene. 

ADD COMMENTlink written 3.1 years ago by yanweng30
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