Characterizing nucleic acid polymorphisms, useful e.g. for detecting inherited disease, by extension of starter primer then single-molecule detection of incorporated nucleotide

摘要

Characterization of nucleic acid (NA) polymorphisms by: (i) attaching at least one starter primer (I) to test NA so that the 3'-end of (I) is downstream of the polymorphism being tested; (ii) extending (I) with at least one fluorescently labeled nucleotide (fnt); and (iii) detecting incorporated fnt by single-molecule detection. Independent claims are also included for the following: (a) similar method, performed in microwells, in which an NA, consisting of single-stranded matrix and (I), is immobilized on a carrier particle ((I) is extended with a fluorescently labeled chain-breaking molecule (II), the wells are (optionally) washed to remove unbound (II) and the label incorporated into (I) is detected); and (b) method for improving efficiency of detection of fluorescence from individual molecules by using a dispersion element to separate lights of different wavelengths.