Detecting breast cancer
Provided herein is technology for breast cancer screening and particularly, but not exclusively, to methods, compositions, and related uses for detecting the presence of breast cancer.
1. A method, comprising:
measuring a methylation level for four genes in a biological sample of a human individual through
treating genomic DNA in the biological sample with a reagent that modifies DNA in a methylation-specific manner;
amplifying the treated genomic DNA using a specific set of primers for each of the selected genes; and
determining the methylation level of the four genes by polymerase chain reaction, nucleic acid sequencing, mass spectrometry, methylation-specific nuclease, mass-based separation, and or target capture;
wherein the four genes are TRIM67, OTX1, MPZ and BANK1.
2. The method of claim 1 , wherein the DNA is treated with a reagent that modifies DNA in a methylation-specific manner.
3. The method of claim 2 , wherein the reagent comprises one or more of a methylation-sensitive restriction enzyme, a methylation-dependent restriction enzyme, and a bisulfite reagent.
4. The method of claim 3 , wherein the DNA is treated with a bisulfite reagent to produce bisulfite-treated DNA.
5. The method of claim 1 , wherein the measuring comprises multiplex amplification.
6. The method of claim 1 , wherein measuring the amount of at least one methylated marker gene comprises using one or more methods selected from the group consisting of methylation-specific PCR, quantitative methylation-specific PCR, methylation-specific DNA restriction enzyme analysis, quantitative bisulfite pyrosequencing, flap endonuclease assay, PCR-flap assay, and bisulfite genomic sequencing PCR.
7. The method of claim 1 , wherein the specific set of primers for each of the four genes is selected from the group consisting of:
for BANK1 a set of primers consisting of SEQ ID NOS: 15 and 16,
for OTX1 a set of primers consisting of SEQ ID NOS: 189 and 190,
for TRIM67 a set of primers consisting of SEQ ID NOS: 247 and 248, and
for MPZ a set of primers consisting of SEQ ID NOS: 175 and 176.
8. The method of claim 1 , wherein the sample comprises tissue.
9. The method of claim 8 , wherein the tissue is breast tissue.
10. The method of claim 1 , wherein the sample is blood, serum, or plasma.