8.5. Second-generation sequencing

Initial genome projects, from small viruses through the human and honey bee project, all relied on ‘Sanger’ dideoxy sequencing, a relatively expensive but accurate protocol developed in the 1980’s that generates sequences (often drawn from random cloned fragments for the popular ‘shotgun’ sequence method) of several hundred to 1000 base pairs. Since 2000, there has been a great economization of sequencing, such that current technologies are more than ten-fold less expensive than Sanger sequencing. Nevertheless, Sanger sequencing persists and is often the right strategy when compared to newer technologies (which currently give either quite short or quite inaccurate sequences). Readers should consider ILLUMINA/SOLEXA sequencing (summarized in the methylation section 9. below), 454 pyrosequencing, SOLiD sequencing or the Ion Torrent platform (all platforms are reviewed by Metzger, 2010), and the final decision might rest on local availability along with different strengths of each platform. As of 2012, most DNA and RNA sequencing efforts include at least some component of ILLUMINA sequencing, as that technology is viewed as being most cost-effective.