Rapid and cost-effective whole exome sequencing for clinical diagnosis and personalized medicine. D. Muzny1, M. Wang1, C. Buhay1, Y. Han1, H. Dinh1, C. Kovar1, H. Doddapanei1, M. Bainbridge1, J. Reid1, E. Boerwinkle1,2, R. Gibbs1 1) Human Genome Sequencing Center, Baylor College of Medicine, Houston, TX 77030; 2) University of Texas Health Science Center at Houston, School of Public Health, Houston, TX 77030.

   For genome sequencing to migrate from the laboratory to clinical care, it should be capable of delivering high-quality sequencing results in a fast, consistent and effective way. A comprehensive approach towards this goal has incorporated protocol development, robotics applications and a robust LIMS so that goals for both high capacity and fast turn around time can be accomplished. To further reduce the turnaround time for whole exome sequencing (WES) in clinical environment, we have developed a fast workflow taking advantage of an experimentally optimized protocol for quick WES library preparation and capture methods as well as shortened sequencing time using the Illumina HiSeq2500 platform. The quick WES library construction protocol allowed us to complete the entire NimbleGen-based whole exome capture library construction within 2 work days (0.5 day for pre-capture library preparation and another 1.5 days for target enrichment), whereas sequencing on HiSeq2500 has helped reduce the data generation time to approximately 30 hours. Kapa HiFi enzyme has been evaluated and utilized in PCR amplification to achieve more uniform target coverage. Validation of the workflow involved multiple co-capture and single-capture tests using benchmark human DNA samples (i.e. HapMap NA12878 and HS1011) and the HGSC-designed VCRome rebalanced probe set (~44Mb capture size). In a recent 5-plex co-capture quick WES experiments, with ~6.0 Gbs of raw data generated for the individual sample, 92% of the total targeted bases were sequenced with a minimum depth of 20 fold. Nearly all (i.e. 99.3%) of the designed VCRome exome targets were present in sequence reads and 98.4% of the targeted bases were covered by at least one read. While the capture efficiency was slightly affected by the short 1-day hybridization scheme, we observed approximately 69% of the reads mapping to the target regions. These experiments indicate that without sacrificing sequencing performance, we are able to reduce the WES turnaround time from the previous three weeks to 3-4 days. Such rapid turnaround times are necessary for WES to be routinely used in prenatal and neonatal intensive care settings.

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