I have searched online for a long time, but there is still no authoritative answer to this question (for single-end sequencing). So, I pointed it out here for clarification for the fresh people using single-end sequencing.
I am using TruSeq Stranded mRNA Sample Prep Kit (a strand-specific kit) to prepare my library. When I sequence (e.g. NovaSeq 6000 sequencing) this library:
- For pair-end sequencing: the first read (read 1) is from the opposite strand (i.e. maps to the antisense strand), and the second read (read 2) is from the transcript strand (i.e. maps to the sense strand).
- For single-end sequencing: all the reads are from the opposite strand (i.e. maps to the antisense strand).
The above info also works for Illumina Stranded mRNA, Illumina Stranded Total, and TruSeq Stranded Total kits, which are all RF/fr-firststrand stranded (dUTP) kits (includingTruSeq Stranded mRNA Kit).
Please feel free to correct me or add any more profound commands or suggestions here. Many thx.