B.S., University of California, Davis, 1984
Ph.D., Baylor College of Medicine, 1996

RESEARCH INTERESTS:
My laboratory is involved in next generation sequencing technology development, which stands to change the way we think about scientific approaches in basic, applied, and clinical research. The broadest application for these next-generation technologies is medical resequencing of human genomes, which could unravel genetic causes of common disease and cancer, assist doctors in prescribing personalized medicine, and provide predictive indicators of disease prior to onset opening the door for preventive therapies. Diabetes mellitus is a complex disease with genetic and environmental components contributing to its onset, progression, and severity. In collaboration with Dr. Ashok Balasubramanyam (Department of Medicine; Division of Diabetes, Endocrinology and Metabolism at BCM), we have begun candidate gene sequencing using the Sanger method, with plans to begin next generation sequencing of diabetic patients prone to ketosis, called ketosis-prone diabetes, in the near future. Of the numerous next generation sequencing strategies under development, my group has focused on the cyclic reversible termination (CRT) method.

CRT is comprised of a three-step process of incorporating modified nucleotides, fluorescence imaging, and deprotecting, after which the cycle begins again. CRT reactions are performed in a high-density format using single molecule arrays or oligonucleotide arrays, eliminating the requirement for gel electrophoresis while significantly increasing sequence throughput. The foundation of this approach is the reversible terminator (RT), its chemical and biological properties of which directly impact the performance of the sequencing technology. Scientists at LaserGen, Inc. have discovered a novel paradigm in RT chemistry, the attachment of a photocleavable, 2-nitrobenzyl group to the nucleobase of dNTP’s, which, upon incorporation, terminates DNA synthesis. The 3’-OH group of the RT remains unblocked, providing favorable incorporation and termination properties for several commercially available DNA polymerases, while maintaining good discrimination against mismatch incorporations. Upon removal of the 2-nitrobenzyl group with UV light, the natural nucleotide is restored without molecular scarring.

We are now at the stage of technology development to begin integration of the CRT chemistry to an instrumentation platform. The key technologies under development are the construction of a microfluidics reaction cell, an imaging system based on our Pulsed-Multiline Excitation (PME) detector technology, and solid-phase CRT sequencing from a high-density, oligonucleotide chip. The CRT system will be integrated with electronic control for reagent delivery to the microfluidics reaction cell, UV illumination and fluorescence imaging, data acquisition, analysis, and storage. Our cost analysis projects an increase in throughput and decrease in cost by approximately two-orders of magnitude, placing the CRT strategy well within the goals of the $100,000 genome.

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SELECTED PUBLICATIONS:
1. Wu W, Stupi BP, Litosh VA, Mansouri D, Farley D, Morris S, Metzker S, Metzker ML (2007). A 3’-OH unblocked nucleotide as a reversible terminator in CRT sequencing, submitted.

2. Metzker ML (2005). Emerging Technologies in DNA Sequencing. Genome Res. 15: 1767-1776.

3. Lewis EK, Haaland WC, Nguyen F, Heller DA, Allen MJ, Macgregor RR, Berger CS, Willingham B, Burns LA, Scott GB, Kittrell C, Johnson BR, Curl RF, Metzker ML (2005). Color-blind fluorescence detection for four-color DNA sequencing. Proc. Natl. Acad. Sci. USA 102: 5346-5351.

4. Gibbs RA, Weinstock GM, Metzker ML, et al. (2004). Genome sequence of the Brown Norway rat yields insights into mammalian evolution. Nature 428: 493 -521.

5. Metzker ML, Mindell DP, Liu, XM, Ptak RG, Gibbs RA, Hillis DM (2002). Molecular Evidence of HIV-1 Transmission in a Criminal Case. Proc. Natl. Acad. Sci. USA 99: 14292-14297.

6. International Human Genome Sequencing Consortium (2001). Initial sequencing and analysis of the human genome. Nature 409: 860-921.

7. Metzker ML, Ansari-Lari MA, Liu XL, Holder DJ, Gibbs RA (1998). Quantitation of Mixed-Base Populations of HIV-1 Variants by Automated DNA Sequencing with BODIPY Dye-Labeled Primers. Biotechniques 25: 446-462.

8. Metzker ML, Lu J, Gibbs RA (1996). Electrophoretically Uniform Fluorescent Dyes for Automated DNA Sequencing. Science 271: 1420-1422.

9. Metzker ML, Raghavachari R, Richards S, Civitello A, Burgess K, Gibbs RA (1994). Termination of DNA synthesis by novel 3'-modified-deoxyribonucleoside 5'-triphosphates. Nucleic Acids Res. 22: 4259-4267.

For more publications, see listing on Pub Med.

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Telephone: 713-798-7565
Fax: 713-798-5741
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Last modified: 6/2007