PYROSEQUENCING-BASED BARCODES FOR DYE-FREE MULTIPLEX BIOASSAY

摘要

We proposed a sequence-based barcoding approach to encode a target with a report base and repeats of two stuffer bases. The barcodes were quantitatively decoded by pyrosequencing without pre-separations, and multiple targets were quantified by peaks of the reporter bases in a pyrogram. Using these barcodes, we have developed a barcoded adapter-based method for multiplex gene-expression analysis; as a proof-of concept, the 12-plex gene expression analysis of four different genes among 12 kinds of tissues of a mouse was successfully demonstrated. In comparison to conventional barcodes based on dyes or nanocrystals, our sequence-based barcodes do not need laser, gel, and electrophoresis. Most importantly, this barcode strategy significantly extends the scope of pyrosequencing capability.