Digital RNA

Digital RNA

Digital RNA sequencing is an approach to RNA-Seq that removes sequence-dependent PCR amplification biases by barcoding the RNA molecules before amplification. RNA is reverse-transcribed to cDNA, and an excess of adapters, each with a unique barcode, is added to the preparation. This barcoded cDNA is amplified and sequenced. Deep sequencing reads are compared, and the barcodes are used to determine the actual distribution of RNA abundance.

Pros:

• Low amplification bias during PCR
• Provides information about abundance of RNA
• Detection of low-copy–number RNA
• Single-copy resolution

Cons:

• Some amplification bias still persists
• Barcodes may miss targets during ligation

• Digital RNA: Shiroguchi K., Jia T. Z., Sims P. A. and Xie X. S. (2012) Digital RNA sequencing minimizes sequence-dependent bias and amplification noise with optimized single-molecule barcodes. Proc Natl Acad Sci U S A 109: 1347-1352

1. This method has been widely integrated into various sequencing techniques due to its high versatility.
2. Digital RNA: Shiroguchi K., Jia T. Z., Sims P. A. and Xie X. S. (2012) Digital RNA sequencing minimizes sequence-dependent bias and amplification noise with optimized single-molecule barcodes. Proc Natl Acad Sci U S A 109: 1347-1352