FIELD: medicine.
SUBSTANCE: BRAF gene fragment under investigation is amplified by asymmetric real-time PCR with two TaqMan probes: BRAF-15-sense - 5'-ROX-aggtgattttggtctagctacagtgaaatc-BHQ2 and BRAF-15-antisense - 5'-Cy5-acccactccatcgagatttcactgta-BHQ2 and primers: direct - 5'-gagatctactgttttcctttactt, reverse - 5'-ggccaaaaatttaatcagt. The stage of primers and probes annealing is 2 s, the number of amplification cycles is 80-100, Taq polymerase concentration is 0.63 units, the annealing temperature of primers and probes is 57°C. Further, the probe-single-stranded DNA duplex melting and construction of melting peaks are performed. Mutations are identified by asymmetry of melting peaks.
EFFECT: method has a high sensitivity, 10 to 15 times higher than the prototype sensitivity, is simple in design, implemented for 2 hours in a single tube with single results recording, reduces the risk of samples cross-contamination.
4 dwg
| Title | Year | Author | Number |
|---|---|---|---|
| METHOD FOR SCANNING GENE MUTATIONS IN HUMAN TUMOURS | 2016 |
|
RU2650862C2 |
| METHOD OF DETECTION OF DELETION IN DAT RAT LINE | 2023 |
|
RU2811140C1 |
| OLIGONUCLEOTIDE SEQUENCE OLIGONUCLEOTIDE SEQUENCE FOR THE NRAS GENE Q61R MUTATION IN THE TUMOR FORMATIONS OF THE THYROID GLAND | 2019 |
|
RU2688189C1 |
| METHOD OF DIAGNOSING MUCOPOLYSACCHARIDOSIS-PLUS SYNDROME | 2022 |
|
RU2798695C1 |
| METHOD FOR ANALYSIS OF SOMATIC MUTATIONS IN BRAF, NRAS AND KIT GENES USING LNA-BLOCKING MULTIPLEX PCR AND SUBSEQUENT HYBRIDIZATION WITH OLIGONUCLEOTIDE BIOLOGICAL MICROCHIP (BIOCHIP) | 2017 |
|
RU2674338C1 |
| METHOD OF ANALYZING TERMINAL MUTATIONS IN BRCA1, BRCA2, ATM AND PALB2 GENES USING MULTIPLEX PCR AND SUBSEQUENT HYBRIDISATION WITH AN OLIGONUCLEOTIDE BIOLOGICAL MICROCHIP (BIOCHIP) | 2020 |
|
RU2729360C1 |
| METHOD FOR ANALYSIS OF SOMATIC MUTATIONS IN GNAQ AND GNA11 GENES USING LNA-BLOCKING MULTIPLEX PCR AND SUBSEQUENT HYBRIDIZATION WITH OLIGONUCLEOTIDE BIOLOGICAL MICROCHIP (BIOCHIP) | 2017 |
|
RU2674687C1 |
| METHOD FOR DETERMINING PULMONARY TUMOUR SENSITIVITY TO TYROSINE KINASE INHIBITOR THERAPY | 2012 |
|
RU2499994C1 |
| METHOD FOR ANALYSIS OF SOMATIC MUTATIONS IN PI3K GENE USING LNA-BLOCKING MULTIPLEX PCR AND SUBSEQUENT HYBRIDISATION WITH OLIGONUCLEOTIDE BIOLOGICAL MICROCHIP (BIOCHIP) | 2013 |
|
RU2549682C1 |
| METHOD FOR DIAGNOSING VARIANT SOPH c.5741G>A IN NBAS GENE | 2024 |
|
RU2819985C1 |
