METHODS AND COMPOSITIONS FOR MODULATION OF AMPLIFICATION EFFICIENCY

摘要

Provided herein are methods and kits for modulating the amplification efficiency of nucleic acids, which are useful in multiplex reactions where the amplification efficiency of one or more nucleic acids in the mixture are desired to be modulated relative to one or more other nucleic acids. Embodiments relate to molecular diagnostics, including detecting sequence variants, such as SNPs, insertions deletions, and altered methylation patterns, as well as the modulation of the amplification efficiency of internal control sequences to provide more accurate control sequences for amplification reactions.

主权项

1. A method to modulate the amplification efficiency of a nucleic acid sequence in an amplification reaction, the method comprising:
providing an amplification reaction comprising a pair of amplification primers comprising a forward primer and a reverse primer, said pair of primers configured to amplify a first target nucleic acid and thereby produce a first amplicon having a first nucleic acid sequence, and configured to amplify a second target nucleic acid and thereby produce a second amplicon having a second nucleic acid sequence, wherein a portion of the second nucleic acid sequence is different from the first nucleic acid sequence; providing a modulator oligonucleotide to the amplification reaction, wherein the modulator oligonucleotide preferentially hybridizes to the second nucleic acid sequence in comparison to the first nucleic acid, and wherein at least a portion of the modulator oligonucleotide shares sequence identity to either the forward or reverse primer, and the remainder of the modulator oligonucleotide hybridizes to at least a portion of the second nucleic acid sequence that differs from the first nucleic acid sequence; amplifying the nucleic acid sequences, wherein the modulator oligonucleotide reduces the amplification efficiency of the second amplicon by competing with at least one of the forward or reverse primer for binding to the second target nucleic acid.