METHOD OF DETECTING STREPTOCOCCUS MUTANS BY ISOTHERMAL LOOP AMPLIFICATION Russian patent published in 2023 - IPC C12Q1/68 

FIELD: medicine; biotechnology; microbiology; molecular biology.

SUBSTANCE: invention relates to the molecular and biological methods for microorganisms detection. A method of detecting Streptococcus mutans by isothermal loop amplification is proposed. 0.12 mM of HNB, Bst DNA polymerase, 1 × reaction buffer (20 mM of Tris-HCl, 10 mM of (NH₄)₂SO₄, 10 mM of KCl, 2 mM of MgSO₄, 0.1% Triton®-X-100, pH 8.8), 5–8 mM of magnesium sulfate, 1.4 mM of deoxynucleotide triphosphates, and a primer set are added to saliva test sample. It is kept at a temperature of 60–65°C for at least 15 minutes, then the presence of Streptococcus mutans DNA in the test sample is detected by changing the color of the reaction mixture from purple to blue. The primer set is selected from the set including primers with the nucleotide sequence Seq Id No: 13–16, 54, a set containing primers with the nucleotide sequence Seq Id No: 19–22, 55, a set containing primers with the nucleotide sequence Seq Id No: 23–26, 58, and a set including primers with the nucleotide sequence Seq Id No: 19–22, 55 and SEQ ID NO: 23–26, 58 and primer count is 1.6 mM for internal primers (FIP, BIP, Seq Id No: 13, 14 or 21, 22 or 23, 24), 0.2 mM for outer primers (F3, B3, Seq Id No: 15, 16, or 19, 20, or 25, 26) and forward loop primer (LF, Seq Id No: 54 or 55 or 58).

EFFECT: invention allows to rapidly detect Streptococcus mutans in a saliva sample without sample preparation with a sensitivity of at least 10-4 ng/μl and a specificity of 100%.

6 cl, 6 dwg, 17 tbl, 5 ex