Method for determining the presence of DNA variants using peptide nucleic acid probes
AssigneeSpectrumedix Corporation
InventorZhaowei Liu
Life of this patent
Estimating the term…
Abstract
The present invention relates to a peptide nucleic acid probe-based method for generating data indicative of the presence of a nucleotide polymorphism, mutation, or methylated cytosine in a nucleotide containing compound. A peptide nucleic acid probe (PNAP) is subjected to temperature gradient electrophoresis in the presence of a nucleotide containing compound. The PNAP is irradiated to generate a spectroscopic signal. The spectroscopic signal is converted into data suitable for determining the presence of the nucleotide polymorphism or the mutation in the nucleotide-containing compound.
Claims23
A peptide nucleic acid probe-based method for generating data indicative of the presence of a variant in a nucleotide containing compound, comprising:
- subjecting a peptide nucleic acid probe (PNAP) having fewer than about 30 bases to temperature gradient electrophoresis in the presence of a nucleotide containing compound;
- irradiating the PNAP to generate a spectroscopic signal;
- and
- converting the spectroscopic signal into data suitable for determining the presence of the variant in the nucleotide-containing compound.
A peptide nucleic acid probe-based method for determining the presence of a variant in a nucleotide containing compound, comprising:
- obtaining a first parameter representative of a first spectroscopic signal resulting from a first peptide nucleic acid probe (PNAP), the first PNAP having been subjected to temperature gradient electrophoresis in the presence of the nucleotide containing compound;
- obtaining a second parameter representative of a second spectroscopic signal resulting from a second PNAP, the second PNAP having been subjected, simultaneously with the first PNAP, to temperature gradient electrophoresis in the presence of the nucleotide containing compound;
- comparing the first and second parameters to determine the presence of the variant in the nucleotide-containing compound.
A peptide nucleic acid probe-based method for determining the presence of variants in a plurality of respective nucleotide containing compounds, comprising:
- subjecting a mixture comprising a plurality of peptide nucleic acid probes and a plurality of different nucleotide containing compounds to temperature gradient electrophoresis, different nucleotide containing compounds having a different length, peptide nucleic acid probes forming complexes with respective nucleotide containing compounds;
- irradiating the peptide nucleic acid probes to generate spectroscopic signals;
- and
- converting the spectroscopic signals into data suitable for determining the presence of variants in respective nucleotide-containing compounds.
A temperature gradient electrophoresis-based method for generating data indicative of the presence of one or more methylated cytosines in a sample comprising a first nucleotide containing compound (NCC) having non-methylated cytosines, comprising:
- contacting the first NCC with a first compound to thereby provide a modified NCC wherein non-methylated cytosines of the first NCC are replaced with a different base;
- amplifying the modified NCC to obtain first PCR products;
- subjecting the first PCR products to temperature gradient electrophoresis in the presence of a peptide nucleic acid probe;
- irradiating the first PCR products and peptide nucleic acid with light to thereby generate a spectroscopic signal;
- and
- converting the spectroscopic signal into data indicative of the presence of the one or more methylated cytosines in the first NCC.
A temperature gradient electrophoresis-based method for generating data indicative of the presence of one or more methylated cytosines in a sample comprising a first nucleotide containing compound (NCC), comprising:
- obtaining first PCR products, the PCR products having been prepared by:
- (a1) contacting the first NCC with a first compound to thereby provide a modified NCC in which non-methylated cytosines are replaced with a different base, and (a2) amplifying the modified NCC to obtain said first PCR products;
- subjecting the first PCR products and a peptide nucleic acid probe (PNAP) to temperature gradient electrophoresis,
- irradiating the first PCR products and peptide nucleic acid (PNAP) with light to thereby generate a spectroscopic signal;
- and
- converting the spectroscopic signal into data indicative of the presence of the one or more methylated cytosines in the first NCC.
A peptide nucleic acid probe-based method for generating data indicative of a sequence of at least a portion of the nucleotide-containing compound, comprising:
- subjecting first and second peptide nucleic acid probes (PNAP's) to temperature gradient electrophoresis in the presence of a nucleotide containing compound, the first and second PNAP's having spectroscopically distinguishable tags;
- irradiating the PNAP's to generate a spectroscopic signal;
- and
- converting the spectroscopic signal into data suitable for determining the sequence of at least a portion of the nucleotide-containing compound.
A peptide nucleic acid probe-based method for generating data indicative of the presence of a variant in a nucleotide containing compound, comprising:
- subjecting a peptide nucleic acid probe (PNAP) to temperature gradient electrophoresis in the presence of a nucleotide containing compound, wherein the nucleotide-containing compound comprises at least about 10 times as many bases as the nucleotide containing compound;
- irradiating the PNAP to generate a spectroscopic signal;
- and
- converting the spectroscopic signal into data suitable for determining the presence of the variant in the nucleotide-containing compound.
Dependencies are inferred from the claim text.