title: Single-nucleotide polymorphism discovery by targeted DNA photocleavage.
abstract: Single-nucleotide polymorphisms are the largest source of genetic variation in humans. We report a method for the discovery of single-nucleotide polymorphisms within genomic DNA. Pooled genomic samples are amplified, denatured, and annealed to generate mismatches at polymorphic DNA sites. Upon photoactivation, these DNA mismatches are then cleaved site-specifically by using a small molecular probe, a bulky metallointercalator, Rhchrysi or Rhphzi. Fluorescent labeling of the cleaved products and separation by capillary electrophoresis permits rapid identification with single-base resolution of the single-nucleotide polymorphism site. This method is remarkably sensitive and minor allele frequencies as low as 5% can be readily detected.
authors.affiliation: Division of Chemistry and Chemical Engineering, California Institute of Technology, Pasadena, CA 91125, USA.
authors.forename: Jonathan R
authors.initials: JR
authors.lastname: Hart
authors.forename: Martin D
authors.initials: MD
authors.lastname: Johnson
authors.forename: Jacqueline K
authors.initials: JK
authors.lastname: Barton
mesh_terms: D000818:Animals; D020137:Base Pair Mismatch / Q000235:genetics; D001483:Base Sequence; D002417:Cattle; D004247:DNA / Q000235:genetics* / Q000528:radiation effects*; D019075:Electrophoresis, Capillary; D005787:Gene Frequency / Q000235:genetics; D005821:Genetic Techniques; D006801:Humans; D008969:Molecular Sequence Data; D009942:Organometallic Compounds / Q000737:chemistry / Q000528:radiation effects; D010782:Photolysis; D010957:Plasmids / Q000235:genetics; D020641:Polymorphism, Single Nucleotide / Q000235:genetics* / Q000528:radiation effects; D011401:Promoter Regions, Genetic / Q000235:genetics / Q000528:radiation effects; D017975:Ruthenium Compounds / Q000737:chemistry / Q000528:radiation effects; D013698:Templates, Genetic; D014409:Tumor Necrosis Factor-alpha / Q000235:genetics / Q000528:radiation effects
chemical_list: D009942:Organometallic Compounds; D017975:Ruthenium Compounds; D014409:Tumor Necrosis Factor-alpha; D004247:DNA