question about unstranded RNA-seq

Question

[biolab]

Hi everyone,

I have an unstranded RNA-seq data. I have a question about this seq type:

if several reads were mapped onto an intergenic region, it indicates presence of a noncoding RNA; however, which strand this ncRNA is derived from? To my understanding, in the unstranded library construction method, both sense and antisense strands are produced before sequencing, so which strand the reads come from?

Any of your answers will be appreciated. Thanks a lot!

Answer 1

[Devon Ryan]

You can't determine which strand a feature is from with an unstranded kit.

[biolab]

Hi Devon,

I make an example as follows. Suppose a read sequence is: 5' AAAAAGGGTTTTT 3'

The two strands are:

• 5' TTTAAAAAGGGTTTTTAAA 3' (+ strand)
• 3' AAATTTTTCCCAAAAATTT 5' (- strand)

So, this read must come from + strand. I can determine the read origin. Am I right? Thanks.

[Brian Bushnell]

You can determine which strand a read maps to, but that does not reflect which strand coded for the RNA, and thus is not particularly useful.

[Parham]

Isn't the strand complementary to the RNA's sequence producing it?

[Nicolas Rosewick]

not in unstranded kit