The draft sequence of the human genome was completed in February 2001, and in the decade since publication sequencing technologies have undergone a revolution in throughput and costs. Current methods employ synthesis or ligation for base-call detection, called ‘2nd generation’ sequencing, while recent improvements have introduced the ‘3rd generation.’ The base-call error-rates vary among the technologies. However, these techniques can generate sequence data rapidly such that an entire genome can be obtained to a reasonable coverage in two to four weeks. Moreover, the number of samples sequenced can be easily increased by using more instruments.
The challenge now facing scientists and physicians is how to best apply the data generated to the clinic. Development of analytical pipelines needed to interpret the sequence data will be critical to its success in clinical application. Among the issues to be solved are the volume of raw data generated and the accuracy of the base-calls. Until the analytical challenges are resolved, the existing sequencing methods remain primarily a research tool, rather than a high-throughput, validated method which could be applied routinely in clinical diagnostics and practice.
Once the analytical challenge is overcome, many of the chip-based methods will be replaced with sequencing that will meet a variety of needs including analysis of genome-wide expression profiling, identification of chromosomal aberrations (i.e. amplification, deletion, translocation and/or inversion), epigenetic modifications and disease-associated mutations. Among the advantages of the approach will be the agnostic nature of the test. Since previous knowledge of targets is not required, this approach allows for better precision of the quantitation and improved ability to discover sequence variations as small as one nucleotide and as large as megabase-sized genomic rearrangements.
Daniela S. Gerhard, Ph.D.
Director, NCI Office of Cancer Genomics