Molecular biological method for rapidly identifying trichosanthin medicinal material
A trichosanthin and molecular technology, applied in the field of molecular identification of trichosanthin and its analogs, can solve the problems of the origin and storage time of medicinal materials, poor repeatability of results, application limitations and the like
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[0005] Three single primers were designed according to the ITS sequence in the trichosanthin ribosomal RNA gene: TkS1-64F, Tks2-1 12F, Tks2-130R. The PCR reaction system is 25 μL, in which Tris-HCl (pH 9.0) 10mmol L -1 , KC1 50mmol·L -1 , Mg 2+ 1.5 mmol·L -1 , each dNTP 0.15 mmol·L -1 , Taq 1U (promega), Primer 0.15 μmol L -1 , template DNA 50 ng. Amplification was carried out on ABI9700. The amplification program was pre-denaturation at 95°C for 4 min, followed by 40 cycles: denaturation at 95°C for 30 s, annealing at 50°C for 45 s, extension at 72°C for 60 s, and extension at 72°C for 5 min after the cycle.
[0006] Among the three primers, TkS1-64F has a good amplification effect, showing that all TCS can be amplified, and most counterfeit products can be amplified, with 1-5 polymorphic bands, clear bands and good repeatability, which can be used for TCS Identification of medicinal materials. The identification bands of trichosanthum were determined to be 560, 960 bp...
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