Assays for short sequence variants

摘要

The invention provides assays that can detect multiple genetic variants of a gene (e.g., a mycobacterium gene) in a sample using a pool (using 2, 3, 4, or more) of oligonucletide hybridization probes.

主权项

1. A kit of reagents capable of identifying which variant among multiple variants of a first region of a gene sequence is present in a sample, said multiple variants differing from one another by one or more changes selected from the group consisting of nucleotide substitutions, deletions, and additions, said kit comprising
a) reagents for amplifying said first region and b) a set of hybridization probes for said first region, said set containing at least three probes that
(i) comprise a single-stranded loop target binding sequence and an arms hybrid,(ii) comprise a nucleotide sequence that is distinct from the other probes, such that each probe has a different target binding sequence,(iii) are detectably differently fluorescently labeled,(iv) hybridize in the presence of said reagents at temperatures in the range of 40-55° C. to more than one variant of said first region with varying degrees of complementarity to form probe-target hybrids having unique melting temperatures, and(v) are separably detectable when hybridized without separating unbound probes wherein said set of hybridized probes is capable through their combined hybridizations to said first region of identifying said variant among multiple variants.