RNA Sequencing Library Preparation

Moving on to RNA sequencing library preparation. In this case, this is usually done to look at gene expression, to identify which genes are turned on or off in a given sample. Most of the times people are really interested in the protein-coding transcripts, or messenger RNAs. These are polyadenylated transcripts. But they only make up a small portion of the total amount of RNA. Most of the RNA is made up of ribosomal RNA, and this is generally things that are undesirable, that people don’t want to sequence and waste their sequencing dollars. To prepare RNA sequencing libraries, there are a couple of extra steps. One is a depletion or an enrichment step. The second is conversion of RNA to DNA, because the Illumina sequencers only work for DNA. You can’t sequence RNA directly on these platforms. And then, the last step, again, is to add sequencing adapters to complete the DNA library preparation of that converted RNA. I’m gonna go over two methods today – the TruSeq library prep method for RNA and also a method called SMART-seq, but there are many, many other types of library preparation for RNA. We just don’t have time to go over all of them in this talk.

Chee-Onn Leong

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