Methods and kits for identifying target nucleotides in mixed populations

Abstract

Ligation-based methods and kits are disclosed for identifying at least two target nucleotides in a mixed population sample that is a sample that contains or potentially contains target nucleic acid sequences from more than one source. Typically, two ligation reaction compositions are formed ligation products generated, and the ligation products or their surrogates are analyzed to identify target nucleotides in the mixed population sample. In certain embodiments, the target nucleic acid sequences, the ligation products, or both are amplified. In certain embodiments, multiplex amplification and/or ligation reactions are performed.

1 . A kit comprising Afu ligase and at least one other thermostable ligase. 
     
     
         2 . The kit of  claim 1 , further comprising at least one first probe set, at least one second probe set, at least one primer, at least one reporter group, at least one affinity tag, at least one hybridization tag, at least one mobility modifier, at least one polymerase, at least one nuclease, or combinations thereof. 
     
     
         3 . The kit of  claim 1 , wherein the at least one other thermostable ligase comprises at least one of: Pfu ligase, Taq ligase,  Thermus  species ligase AK16D, Tth ligase, Tsc ligase, Tfi ligase, Mth ligase, Ape ligase, TS2126 ligase, lig Tk  ligase, or combinations thereof. 
     
     
         4 . The kit of  claim 1 , further comprising at least one first probe set, at least one second probe set, at least one primer, at least one reporter group, at least one affinity tag, at least one hybridization tag, at least one mobility modifier, at least one polymerase, at least one nuclease, or combinations thereof.