Patent ID: 8129120
Filing Date: 2012-03-06
Classification: C12Q

Abstract:
1. A method for determining the nucleotide present at a selected polymorphic site in a target nucleic acid molecule, the method comprising: a) amplifying a portion of the target nucleic acid molecule comprising the selected polymorphic site using a first primer and a second primer, the second primer containing a recognition site for a first restriction enzyme and a recognition site for a second restriction enzyme, to generate amplification product containing a recognition site for the first restriction enzyme and a recognition site for the second restriction enzyme such that digestion of the amplification product with the first restriction enzyme and the second restriction enzyme generates a nucleic acid fragment containing the selected polymorphic site; b) digesting the amplification product with the first restriction enzyme and the second restriction enzyme to generate a nucleic acid fragment containing the selected polymorphic site within a 5′ single-stranded overhang; c) filling the recessed end corresponding to the 5′ overhang with nucleotides wherein at least one of the nucleotides is bromodeoxuridine; and d) analyzing the nucleic acid fragment to identify the nucleotide present at the selected polymorphic site.