Hybrid approach to DNA sequencing

The traditional Sanger method of DNA sequencing, which uses dideoxynucleotides (ddNTPs) to terminate DNA synthesis at each of the four bases, has remained the gold standard in DNA sequencing for years. The recent development of ‘‘sequencing by synthesis’’ (SBS) techniques, which identify each base as the DNA strand is extended, has offered faster and cheaper sequencing in a massively parallel scale. Jia Guo et al. report an alternative DNA sequencing technique that combines the advantages of Sanger with the high-throughput capability of SBS methods. The authors modified the four nucleotides by capping the 3 -OH end with a small, reversible moiety that still allows DNA polymerase to recognize the modified nucleotides, thus creating ‘‘reversible terminators.’’ They then combined these nucleotides with cleavable fluorescent ddNTPs. As the extension cycles proceed, the ratio of the two nucleotide sets is adjusted, and the sequences are determined by the unique fluorescence emission of the DNA products terminated by the ddNTPs. This hybrid four-color sequencing approach on a DNA chip—which accurately sequenced DNA templates with read lengths 30 bases—has potential application in ultra-highthroughput DNA sequencing, the authors say. — M.M.