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Paired-end RNA sequence - which read is complimentary?

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 next-gen sequence gene
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updated by Ram Ram • written by yanweng • yanweng

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

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updated by Ram Ram • written by yanweng • yanweng

2 answers

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

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written by Antonio R. Franco Antonio R. Franco

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.

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written by Devon Ryan Devon Ryan

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?

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written by apelin20 apelin20

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