DNA barcode based premier for identifying garter snake, PCR-RFLP method, and kit

Abstract

The invention belongs to the technical field of traditional Chinese medicine species identification, and provides a polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP) method for identifying garter snake. The sequence of the provided PCR amplification primer is DK1-CO1:5'-CAA CTA ACC ACA AAG ACA TCG G-3', DK1-CO2:5'-CTT CTG GGT GGC CGA AAATC A-3'; the characteristic DNA base sequences of garter snake are 5'-ACTAGT-3', which is positioned at the 121st position to 126th position of the COI sequence, and 5'-GGAAACC-3', which is positioned at the 353rd position to the 359th position of the COI sequence; the restriction endonuclease Spe I can identify and cut the characteristic sequence 5'-GGAAACC-3', the restriction endonuclease BstE II cannot identify the characteristic sequence 5'-GGAAACC-3', and the two restriction endonucleases can only cut garter snake double-enzyme into two segments. The identification process comprises the following steps: extracting the DNA of a sample, using primers DK1-CO1 and DK1-CO2 to carry out PCR amplification, purifying the PCR products, using restriction endonucleases Spe I and BstE II to digest the PCR products; carrying out agarose gel electrophoresis to obtain the analysis result, comparing the analysis result with the PCR-RFLP characteristic result of garter snake to determine whether the provided sample is garter snake or not. The invention also provides a kit for the identification method. The identification method is accurate, rapid, and reliable, and can be widely applied to identification of traditional Chinese herbals.

Description

technical field [0001] The invention relates to the technical field of variety identification of traditional Chinese medicines, in particular to a primer for identifying black snakes, a PCR-RFLP identification method and a kit. Background technique [0002] Zaocys dhumnades is derived from the dried body of snake snakes. It has the effects of expelling wind, dredging collaterals, and relieving spasm. At present, there are more and more confused varieties of black-snake medicinal materials in commercial circulation, and the common species of the snake family may be regarded as black-snake. difficulty. Therefore, finding a fast and accurate method to identify black snake is an important task to ensure the safety of medicine for black snake. Contents of the invention [0003] The object of the present invention is to provide a kind of PCR-RFLP molecular identification method of accurately distinguishing black snake, comprising the characteristic DNA base sequence of black s...

AI Technical Summary

Problems solved by technology

At present, there are more and more confused varieties of black-snake medicinal materials in commercial circulation, and the common species of the snake family may be regarded as black-snake. difficulty

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