An integrated system for DNA sequencing by synthesis using novel nucleotide analogues

The Human Genome Project has concluded, but its suc-cessful completion has increased, rather than decreased, the need for high-throughput DNA sequencing technologies. The possibility of clinically screening a full genome for an individual’s mutations offers tremendous benefits, both for pursuing personalized medicine and for uncovering the genomic contributions to diseases. The Sanger sequencing method, although enormously productive for more than 30 years, requires an electrophoretic separation step that, unfor-tunately, remains a key technical obstacle for achieving eco-nomically acceptable full-genome results. Alternative sequencing approaches thus focus on innovations that can reduce costs. The DNA sequencing by synthesis (SBS) approach has shown great promise as a new sequencing platform, with par-ticular progress reported recently. The general fluorescent SBS approach involves (i) incorporation of nucleotide analogs bear-ing fluorescent reporters, (ii) identification of the incorporated nucleotide by its fluorescent emissions, and (iii) cleavage of the fluorophore, along with the reinitiation of the polymerase reaction for continuing sequence determination. In this Account, we review the construction of a DNA-immobilized chip and the development of novel nucleotide reporters for the SB