End-capped HyBeacon probes for the analysis of human genetic polymorphisms related to warfarin metabolism.
HyBeacon probes have been used to characterise SNPs in the CYP2C9 and VKORC1 genes associated with variations in the efficiency of warfarin metabolism. PCR amplification of genomic DNA and probe target melting analysis provided a robust and reliable method to differentiate polymorphic sequences at these loci. Probes capped with 5'-trimethoxystilbene were found to exhibit larger fluorescence differences between hybridised and dissociated states than uncapped probes, generating melting peaks of considerably improved height. 3'-Pyrene modifications enhanced probe signal-to-noise and melting peak heights with the additional benefit of acting as PCR stoppers, and 5',3'-doubly capped probes gave the best analytical results.
Ben Gaied
Nouha
N
School of Chemistry, University of Southampton, Highfield, Southampton, UKSO17 1BJ.
Richardson
James A
JA
Singleton
Daniel G
DG
Zhao
Zhengyun
Z
French
David
D
Brown
Tom
T
D001189:Aryl Hydrocarbon Hydroxylases / Q000235:genetics; D065729:Cytochrome P-450 CYP2C9; D004247:DNA / Q000235:genetics / Q000378:metabolism; D005456:Fluorescent Dyes / Q000737:chemistry*; D005838:Genotype; D006801:Humans; D006899:Mixed Function Oxygenases / Q000235:genetics; D015345:Oligonucleotide Probes / Q000737:chemistry*; D016133:Polymerase Chain Reaction / Q000379:methods; D011110:Polymorphism, Genetic; D064417:Vitamin K Epoxide Reductases; D014859:Warfarin / Q000378:metabolism*
D005456:Fluorescent Dyes; D015345:Oligonucleotide Probes; D014859:Warfarin; D004247:DNA; D006899:Mixed Function Oxygenases; C450260:CYP2C9 protein, human; D065729:Cytochrome P-450 CYP2C9; D001189:Aryl Hydrocarbon Hydroxylases; C578523:VKORC1 protein, human; D064417:Vitamin K Epoxide Reductases