Molecular marker for identifying common threewingnut root medicinal material, primer and identification method
A technology of molecular markers and Tripterygium wilfordii, which is applied in biochemical equipment and methods, microbial measurement/testing, DNA/RNA fragments, etc., can solve the problems of strong subjectivity and poor repeatability, and achieve high sensitivity, strong specificity, highly reproducible results
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Embodiment 1
[0056] This example provides molecular markers and primer pairs for identifying the authenticity of Tripterygium wilfordii.
[0057] Download the nucleotide sequences of Tripterygium wilfordii and its common counterfeit products from the NCBI database. See Table 2 for the names and GenBank accession numbers. Using BioEdit software, for the gene sequences of all downloaded Tripterygium wilfordii and its counterfeit products, the comparison analysis of psbA-trnH, rbcL, ITS and matK sequences was carried out. There are no stable SNP variation sites between species and matK sequences, and there are SNP variation sites within species. Therefore, the above three sequences cannot be used for inter-species identification, and there are a large number of repetitive sequences in the psbA-trnH gene sequence itself. Suitable for PCR primer design. However, through the analysis, it was found that there were many variation sites in the ITS sequence of the genuine Tripterygium wilfordii and...
Embodiment 2
[0065]This example provides a PCR reagent for identification of tripterygium wilfordii, including a pair of primers LGTITS-715A (primer 1) and LGTITS-263A (primer 2), DNA polymerase and dNTPs.
[0066] Further, the PCR reagents include:
[0067] 2×Taqplus Master Mix II, 25μL;
[0068] Primer 1, the concentration is 6.4ng / μL, 2μL;
[0069] Primer 2, the concentration is 6.4ng / μL, 2μL;
[0070] DNA template, 200-800ng;
[0071] Add sterilized double distilled water to 50 μL.
Embodiment 3
[0073] This embodiment provides a kit for identification of tripterygium wilfordii, including the PCR reagent in embodiment 2.
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