Next-generation sequencing: from basic research to diagnostics.
BACKGROUND
For the past 30 years, the Sanger method has been the dominant approach and gold standard for DNA sequencing. The commercial launch of the first massively parallel pyrosequencing platform in 2005 ushered in the new era of high-throughput genomic analysis now referred to as next-generation sequencing (NGS).


BACKGROUND
This review describes fundamental principles of commercially available NGS platforms. Although the platforms differ in their engineering configurations and sequencing chemistries, they share a technical paradigm in that sequencing of spatially separated, clonally amplified DNA templates or single DNA molecules is performed in a flow cell in a massively parallel manner. Through iterative cycles of polymerase-mediated nucleotide extensions or, in one approach, through successive oligonucleotide ligations, sequence outputs in the range of hundreds of megabases to gigabases are now obtained routinely. Highlighted in this review are the impact of NGS on basic research, bioinformatics considerations, and translation of this technology into clinical diagnostics. Also presented is a view into future technologies, including real-time single-molecule DNA sequencing and nanopore-based sequencing.


CONCLUSIONS
In the relatively short time frame since 2005, NGS has fundamentally altered genomics research and allowed investigators to conduct experiments that were previously not technically feasible or affordable. The various technologies that constitute this new paradigm continue to evolve, and further improvements in technology robustness and process streamlining will pave the path for translation into clinical diagnostics.
Voelkerding
Karl V
KV
ARUP Institute for Experimental and Clinical Pathology, Salt Lake City, Utah 84108, USA. voelkek@aruplab.com
Dames
Shale A
SA
Durtschi
Jacob D
JD
D000818:Animals; D001483:Base Sequence / Q000235:genetics*; D035843:Biomedical Research / Q000379:methods*; D023281:Genomics; D006801:Humans; D025202:Molecular Diagnostic Techniques / Q000379:methods*; D017712:Peer Review, Research / Q000379:methods*; D012984:Software