Herein we propose to further develop and utilize an ultra-high throughput DNA sequencing technology to generate large amounts of genome sequence to identify and characterize genomic variation in a population of people diagnosed with invasive melanoma. Specifically, we will ultimately sequence chromosome 6 from ten carefully chosen individuals. As a quantitative milestone, we will report the sequence for chromosome 6 to at least 85% completion for these ten people at better than 99.999% assembled accuracy. In a future R33 proposal, we will collect sequence for 290 additional people. In the end, we will have a robust platform for generating high-throughput nucleic acid sequence and converting these data to genetic markers that identify people who are predisposed to progressive malignant melanoma. Current progress towards our goals is at an advanced stage. We are able to routinely sequence bacterial genomes and we have the potential to generate enough data to rapidly sequence a single human chromosome >20x coverage in less than a month. Therefore, we feel that much of the technical risk has been reduced by our previous work, and there are substantial rewards to be gained by pursuing the goals described here.
| Xu, Ming Yan; Aragon, Anthony D; Mascarenas, Monica R et al. (2010) Dual primer emulsion PCR for next- generation DNA sequencing. Biotechniques 48:409-12 |
