Nextera XT on RNA seq?

[helico93561]

Hi all,

I am planning to do an expression study by sequencing mRNA.

The standard protocol would be following Truseq sample prep kit.

However, I read somewhere that
1. remove riboRNA by Ribo Zero
2. add poly A tail to leftover RNA
3. use poly dT for cDNA synthesis

If that's the case, is it possible to use Nextera or Nextera XT to sequence the cDNA?

[mcnelson.phd]

The simple answer is that yes, you can use Nextera/XT on your cDNA to make a pretty decent and quick RNASeq library.

However, you will lose the 5' and 3' ends, which can bias some analyses. There will also be some other sources of bias in the library that can affect your results when using Nextera such as longer transcripts having more representation while very short transcripts will not be represented at all. You also won't have any strand information which means you won't be able to look for regulatory RNAs.

Also, you'll want to consider where you'll be sequencing and on what instrument. Not all cores will take Nextera libraries to run on a HiSeq, although that's probably less common than it was a year ago.

[Dimitri Szymczak]

The most similar solution to Nextera for RNA-seq is the ScriptSeq.
It was developped by the same manufacturer.