Nanopore sequencing of DNA with MspA

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DESCRIPTION (provided by applicant): The objective of this renewal proposal is to bring nanopore sequencing technology with MspA to fruition. With funding through this program, our group has engineered the porin MspA specifically for nanopore sequencing. By combining MspA with a processive enzyme technology our team demonstrated the visionary goal of nanopore sequencing first conceived two decades ago. With continued funding, we will take the necessary steps to ready nanopore sequencing for industrial integration. We will also explore improvements and variations that make nanopore sequencing an even more powerful tool. Our specific aims are: (1) develop the base calling algorithms necessary for our present nanopore sequencing process, (2) further refine the porin MspA through rational - and bold - genetic engineering in order to optimize its sequencing quality, (3) understand and improve DNA translocation dynamics by reducing its stochastic character and (4) demonstrate sequencing of long segments of native genomic DNA including identification of epigenetic modifications. Our team's success to date has enabled us to form partnerships and gain support from many excellent labs whose expertise has assisted us to move nanopore sequencing forward. We will work with our partner labs to complete the aims outlined in this proposal. It is our goal to improve the quality of human life by delivering a technique that can accurately sequence a human genome for well under $1000 and provide sequencing results within the short timeframe required for clinical practice.