| 摘要 |
FIELD: medicine. ^ SUBSTANCE: method involves the annealing of a specific DNA-primer on an analysed RNA matrix, the reverse transcriptase elongation of the primer in a buffer mixture in the presence of specific terminators and the analysis of reverse transcription products. Reverse transcriptase is presented by a recombinant form of reverse transcriptase of a Maloney murine leukaemia virus (MMLV) being a protein product of the truncated gene pol of the MMLV virus, containing a DNA-polymerase domain, but free from a ribonuclease domain in the concentration of 3-6.7 e.a./mcl. The specific terminators are proper dideoxyribonucleoside triphosphates (ddNTP). The final concentration of dNTP relevant to the present ddNTP makes 20-30 mcM, of the others dideoxyribonucleoside triphosphates - 80-130 mcM. What is applied as the buffer mixture is a mixture containing 50 mMKCI, 50 mM Tris-HCl (pH 8.3 at 25C), 10 mM DTT, 4 mm MgCl2, 1 mM water-soluble salt Mn (II). The primer is elongated for 30-120 min at 37-45 C that is followed by the gel electrophoresis analysis of the reverse transcription products. ^ EFFECT: invention enables simplifying and reducing the price of the method for determining the primary structure of RNA with high specificity preserved. ^ 3 cl, 3 dwg, 3 ex |
