Research:
High-throughput technologies are having a transformative impact on biomedical research. We are broadly interested in developing new experimental methods and computational tools for the parallelized interrogation of biological systems. Ongoing projects in the laboratory make extensive use of new platforms for array-based, programmable DNA synthesis and massively parallelized, short-read DNA sequencing. Areas of active interest include:
Genome partitioning & targeted variation discovery. Second-generation DNA sequencing technologies have the potential to markedly accelerate genetics research, but are hindered by the lack of equivalently powerful methods to selectively isolate complex, megabase-scale subsets of a mammalian genome (e.g. the discontiguous ~31 megabases of protein-coding subsequences, or a contiguous 3 megabase region to which a phenotype has been mapped). We have therefore developed aqueous-phase and solid-phase protocols for the multiplex capture of complex, arbitrary genomic subsets. We have coupled these methods to second-generation sequencing platforms to achieve sensitive and specific variation discovery within targeted regions. Current work is focused on further develo ping , optimizing and applying these strategies to achieve cost-effective resequencing of targeted regions within individual human and mouse genomes. We are particularly interested in develo ping a method that enables the specific and uniform capture and resequencing of the ~180,000 subsequences (aggregate of 31 Mb) that collectively comprise the ‘protein-coding genome’.
High-throughput screening & functional dissection of cis-regulatory elements (“CREs”). For all but a few well-studied examples, we lack quantitative, predictive models for defining how CREs (e.g. enhancers and promoters) integrate variable inputs to yield specific levels of gene activation. In addition, the annotation of functional CREs in mammalian genomes is far from complete. We are developing a platform for high-throughput screening and high-resolution functional analysis of CREs. Our approach relies on the parallel, cost-effective generation of thousands of synthetic enhancers or promoters that are linked in cis to transcribed barcode sequences. After assaying these constructs in parallel, deep sequencing of the barcodes present within cDNA is used to estimate the functional activity of the individual synthetic promoters to which the barcodes were linked. We have successfully demonstrated this approach on well-characterized bacteriophage promoters, and are presently transitioning to more complex systems.
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Publications:
Porreca GJ, Zhang K, Li JB, Xie B, Austin D, Vassallo SL, LeProust EM, Peck BJ, Emig CJ, Dahl F, Gao Y, Church GM, Shendure J. Multiplex amplification of large sets of human exons. Nat Methods. 2007 Nov;4(11):931-6. Epub 2007 Oct 14.
Shendure J*, Porreca GJ*, Reppas NB, Lin X, McCutcheon JP, Rosenbaum AM, Wang MD, Zhang K, Mitra RD, Church GM. Accurate multiplex polony sequencing of an evolved bacterial genome. Science. 2005 Sep 9;309(5741):1728-32. Epub 2005 Aug 4.
Shendure J, Mitra RD, Varma C, Church GM. Advanced sequencing technologies: methods and goals. Nat Rev Genet. 2004 May;5(5):335-44. Mitra RD, Shendure J, Olejnik J, Edyta-Krzymanska-Olejnik, Church GM. Fluorescent in situ sequencing on polymerase colonies. Anal Biochem. 2003 Sep 1;320(1):55-65.
Zhu J*, Shendure J*, Mitra RD, Church GM. Single molecule profiling of alternative pre-mRNA splicing. Science. 2003 Aug 8;301(5634):836-8.
Shendure J, Church GM. Computational discovery of sense-antisense transcription in the human and mouse genomes. Genome Biol. 2002 Aug 22;3(9):RESEARCH0044. Epub 2002 Aug 22.
Weber G*, Shendure J*, Tanenbaum DM, Church GM, Meyerson M. Identification of foreign gene sequences by transcript filtering against the human genome. Nat Genet. 2002 Feb;30(2):141-2. Epub 2002 Jan 14.
Aach J*, Bulyk ML, Church GM, Comander J, Derti A, Shendure J*. Computational comparison of two draft sequences of the human genome. Nature. 2001 Feb 15;409(6822):856-9.
additional publication listings available via PubMed
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Office Phone: (206) 685-8543