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 four 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 target capture;
wherein the four genes are ITPRIPL1, FAM59B, TRH_A and c17orf64_B.
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 selected genes is selected from the group consisting of:
for TRH_A a set of primers consisting of SEQ ID NOS: 245 and 246,
for ITPRIPL1 a set of primers selected from the group consisting of from SEQ ID NOS: 97 and 98, SEQ ID NOS: 99 and 100 and SEQ ID NOS: 425 and 426,
C17orf64_B a set of primers selected from the group consisting of SEQ ID NOS: 269 and 270 and SEQ ID NOS: 449 and 450, and
for FAM59_B a set of primers consisting of SEQ ID NOS: 427 and 428.
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.