14.3: The Biology of mRNA Sequencing

The first step in mRNA sequencing is to lyse the cells of interest. This creates a mass of proteins, nucleotides, and other molecules which are then filtered through so that only RNA (or specifically mRNA) molecules remain. The resulting transcripts are then fragmented into reads 200-1000 base pairs long and undergo a reverse transcription reaction to build a strand-specific DNA library. Finally, both ends of these DNA fragments are sequenced. After establishing these sequenced reads, the computational part of RNA-Seq can be divided into three parts: read mapping, reconstruction, and quantification.

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