Suppression of non-specific amplification with high-homology oligonucleotides

摘要

The invention comprises suppressor oligonucleotides for reducing amplification of a non-target nucleic acid sequences; the method of designing and using such oligonucleotides, as well as kits and reaction mixtures.

主权项

1. A reaction mixture for performing an amplification reaction of a target sequence in the genome of a target organism with reduced amplification of non-target sequences, comprising:
at least one allele-specific primer specific for a variant of the target sequence; an amplicon formed by the at least on allele-specific primer and a reverse primer; a suppressor oligonucleotide (a) having no significant similarity with the target sequence as determined by the blastn algorithm for somewhat similar sequences, (b) extendable by a DNA polymerase, and (c) having a sequence comprising at least one region of complementarity to multiple sites in the genome of the target organism, wherein the least one region of complementarity: (i) is at least 15 base pairs long; and (ii) spans the 3′-end of the suppressor oligonucleotide; and a fluorescently labeled probe oligonucleotide, wherein the reaction mixture does not include an amplicon formed by the suppressor oligonucleotide.