Personal Genomes & Medicine. HST Advisory Council Thursday 16-Nov-2004 2:00 to 2:20 PM. Thanks to: Broad Inst., DARPA-BioComp, DOE-GTL , EU-MolTools, NGHRI-CEGS , NHLBI-PGA, NIGMS-CECBSR, PhRMA, Lipper Foundation
HST Advisory Council
Thursday 16-Nov-2004 2:00 to 2:20 PM
Broad Inst.,DARPA-BioComp, DOE-GTL,EU-MolTools,
NGHRI-CEGS, NHLBI-PGA, NIGMS-CECBSR,PhRMA, Lipper Foundation
Agencourt, Ambergen, Atactic, BeyondGenomics, Caliper, Genomatica, Genovoxx, Helicos, MJR, NEN, Nimblegen, ThermoFinnigan, Xeotron/Invitrogen
For more info see: arep.med.harvard.edu
Shendure et al. 2004 Nature Rev Gen 5, 335.
Do association studies need the added baggage of "linkage" assumptions?
Should we determine genotype (haplotype) directly (at low cost) rather than infer it from population trends?
Even "dispensable" regions of the genome can harbor neomorphic alleles. Each of us has about 104 mutations since the last major population bottleneck.
"-463GA, has been associated with incidence or severity of inflammatory diseases, including atherosclerosis and Alzheimer's disease, and some cancers. The polymorphism is within an Alu element " Kumar AP, et al. (2004) J Biol Chem. 279:8300-15.
Variable breakpoints in Burkitt lymphoma cells with chromosomal t(8;14) translocation separate c-myc and the IgH locus up to several hundred kb. Joos S, et al. (1992) Hum Mol Genet. 1:625-32.
Multiple rare alleles contribute to low plasma levels of HDL cholesterol. Cohen JC et al. (2004) Science. 305:869-72.
Mutations G719S, L858R, Del746ELREA in red.
EGFR Mutations in lung cancer: correlation with clinical response to gefitinib [Iressa] therapy. Paez, … Meyerson (2004) Science 304: 1497
Dulbecco R. (1986) A turning point in cancer research: sequencing the human genome. Science 231:1055-6.
(1) Single-cells: Preimplantation (PGD), uncultivatable
(2) Co-occurrence on a molecule, complex, cell
RNA splice-forms & DNA haplotypes
(3) Cost: $1K-100K "personal genomes"
(4) Precision: Counting 109 RNA tags (to reduce variance)
(~5e5RNAs per human cell)
Fixed 5e3 5e4 5e6 5e9 (goal)
costs EST SAGE MPSS Polony-FISSeq
Zhu,J, et al. Science. 301:836-8.
Eph4 = murine mammary epithelial cell line
Eph4bDD = stable transfection of Eph4 with MEK-1 (tumorigenic)
Zhu J, Shendure J, Mitra RD, Church GM. Science 301:836-8.
Single molecule profiling of alternative pre-mRNA splicing.
Throughput = (# loci × # samples) / time
Monolayered immobilization in acrylamide
Images → Tag Sequences
Tag Sequences → Genome
Bead Polony Sequencing Pipeline
Bead polonies via
Enrichment of amplified beads
FISSEQ or “wobble”
Epifluorescence Scope with Integrated Flow Cell
Mitra,RD, Shendure,J, Olejnik,J, Olejnik,EK, and Church,GM (2003) Fluorescent in situ Sequencing on Polymerase Colonies. Analyt. Biochem. 320:55-65
Cy5 primer (570nm) ; Cy3 dNTP (666nm)
Self Organizing Monolayer
Required disclosure > optional > required privacy