SET OF OLIGONUCLEOTIDE PRIMERS AND PROBES FOR DETERMINING ALLELES OF THE RS55986091 POLYMORPHISM AND METHOD FOR ITS USE Russian patent published in 2023 - IPC C12Q1/68 

FIELD: biotechnology.

SUBSTANCE: set of oligonucleotide primers and probes has been proposed to determine alleles of the rs55986091 polymorphism, which is associated with susceptibility to cervical cancer. The kit includes direct primer 6091-F - SEQ ID NO: 1; TCT TTT TTA TTT TTC CTA AGA GTC GAT, reverse primer 6091-R - SEQ ID NO: 2; GCA ATT TTA ATT TCG CCT CAG TC, Fluorescence Probe 6091-G- SEQ ID NO: 3. (FAM)TTA TTA GGG ATT TTC A (BHQ1), fluorescent probe 6091-A - SEHQ ID NO: 4. (R6G)TTA TTA GAG ATT TTC AA (BHQ1). A method has been proposed for using a set of oligonucleotide primers and probes to determine the alleles of the rs55986091 polymorphism. The method involves extracting DNA from biological material and performing PCR-RRV, amplification, analysis and interpretation of the results. The presence of nucleotide G in a biological sample is determined by assessing the accumulation curves of fluorescent signals along the channel for the FAM fluorophore. The presence of nucleotide A in a biological material sample is determined by assessing the accumulation curves of fluorescent signals along the channel for the R6G fluorophore. The result obtained is interpreted as follows: (i) a sample of biological material has the GG genotype, provided that when examined using the FAM fluorophore channel, a Ct value of 37 or less is determined, and the R6G fluorophore channel does not have a Ct value or there is a difference between the Ct values for channels for fluorophores R6G and FAM more than 2; (ii) a sample of biological material has the AA genotype if, when examined using the R6G fluorophore channel, a Ct value of 37 or less is determined, and there is no Ct value in the FAM fluorophore channel or the difference between the Ct values in the FAM and R6G fluorophore channels is greater than 2; (iii) a sample of biological material has the GA genotype if, during its examination, the channels for the FAM and R6G fluorophores are determined to have a Ct equal to or less than 37, and the difference in Ct absolute values between the channels for the FAM and R6G fluorophores is less than 2.

EFFECT: synthesized oligonucleotide primers SEQ ID NO: 1-2 allow amplification of a fragment between the HLA-DQA1 and HLA-DQB1 genes and do not cross-react with other genome sequences; synthesized oligonucleotide probes SEQ ID NO: 3-4 make it possible to determine alleles of the rs55986091 polymorphism (substitution G>A) and detect the genotype with 100% specificity.

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