Question: Is illumina small RNA-Seq strand specific?
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gravatar for jil340
4.3 years ago by
jil3400
United States
jil3400 wrote:

Hi,

I just wonder if illumina small RNA-Seq is strand specific, that is, if a read is mapped on forward strand, it comes exactly from the forward strand, and vice versa. 

The illumina TruSeq small RNA Sample Prep Guide says "The RNA 3' adapter is specifically modified to target microRNAs and other small RNAs that have a 3' hydroxyl group resulting from enzymatic cleavage by Dicer or other RNA processing enzymes".  I just wonder if the sequence of 3' and 5' adaptor are different, and if the strand information is preserved in the process.

Thanks a lot!

sequencing rna-seq • 3.8k views
ADD COMMENTlink modified 4.3 years ago by jon.brate200 • written 4.3 years ago by jil3400

I think that would be also depending on the way library was prepared (if its strand specific). 

Once you have sequencing data then the mapped file can be checked on RNA-Seq QC level for strandness info.

ADD REPLYlink modified 4.3 years ago • written 4.3 years ago by Manvendra Singh2.0k

You can use "infer_experiment.py" script, available in RSeQC package (http://rseqc.sourceforge.net/#infer-experiment-py), to check how reads were stranded.

ADD REPLYlink written 4.3 years ago by iraun3.5k
0
gravatar for jon.brate
4.3 years ago by
jon.brate200
Norway
jon.brate200 wrote:

I think that most smallRNA-libraries are strand specific because they usually attach an adaptor (or a poly-A tail) prior to fragment amplification. And as long as something is attached to either end of a fragment prior to amplification you retain strand-information.

ADD COMMENTlink written 4.3 years ago by jon.brate200

So if it is strand specific, which strand should I expect the reads to be aligned to if I try to map reads to precursor sequences downloaded from miRbase? I am using illumina small rna lib prep kit, so should the reads be mapped to forward strand of the miRNA (as provided in the precursor fasta file) or its reverse complementary?

ADD REPLYlink written 2.5 years ago by gujh0
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