METHOD, COMPOSITION AND KIT FOR FLUORESCENT QUANTITATIVE PCR AND THEIR APPLICATION Russian patent published in 2023 - IPC C12Q1/686 

FIELD: biotechnology.

SUBSTANCE: invention relates to biotechnology. A method of fluorescent quantitative PCR is described. Pairs of direct and reverse primers, a fluorescent probe and a reagent for PCR amplification are mixed. Fluorescent quantitative PCR is carried out. In this case, the fluorescent probe has two quenching groups, where the first quenching group is located at the 3'-end, and the second quenching group is attached on the basis of T and is located at a distance of 10-15 nucleotides from the first quenching group. An additional additive is added to the mixture, and an additional additive is any one or more of the formamide, DSN and antibiotics Proclin. The method of single-stage fluorescent quantitative PCR is disclosed. A sample release agent, an additional additive, pairs of direct and reverse primers, a fluorescent probe, a reagent for PCR amplification and a sample are mixed. Fluorescent quantitative PCR is carried out. In this case, the fluorescent probe has two quenching groups, where the first quenching group is located at the 3'-end, and the second quenching group is attached on the basis of T and is located at a distance of 10-15 nucleotides from the first quenching group. The method does not include steps of purification and /or extraction of nucleic acid. An additional supplement is any one or more of the formamide, DSN and Proclin antibiotics. The composition is presented for fluorescent quantitative PCR, containing a pair of forward and reverse primers, a fluorescent probe and a reagent for PCR amplification, where the fluorescent probe has two quenching groups, where the first quenching group is located at the 3'-end, and the second quenching group is attached on the basis of T and is located at a distance of 10-15 nucleotides from the first quenching group, where the composition contains an additional additive, which is any one or more of formamide, DSN and Proclin antibiotics. A set for fluorescent quantitative PCR containing the specified composition is also presented.

EFFECT: invention makes it possible to improve the sensitivity of PCR and reduce the number of false negative results.

6 cl, 6 dwg, 12 tbl, 5 ex