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ITS sequence, specific primer pair, identification kit, and identification method used for identification of dendrobium officinale and high imitation fake products

A technology of Dendrobium officinale and specificity, applied in the field of molecular biology, can solve the problem of inability to accurately distinguish Dendrobium officinale, and achieve the effects of simple operation, good specificity and high sensitivity

Inactive Publication Date: 2018-12-07
INFINITUS (CHINA) CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, there are still many species similar to Dendrobium candidum with the ITS2 gene sequence highly similar to Dendrobium candidum, so it is impossible to accurately distinguish Dendrobium candidum from its counterfeit products

Method used

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  • ITS sequence, specific primer pair, identification kit, and identification method used for identification of dendrobium officinale and high imitation fake products
  • ITS sequence, specific primer pair, identification kit, and identification method used for identification of dendrobium officinale and high imitation fake products
  • ITS sequence, specific primer pair, identification kit, and identification method used for identification of dendrobium officinale and high imitation fake products

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] Embodiment 1, be used to detect the synthesis of the PCR primer of Dendrobium officinale

[0038] A total of 170 ITS sequences of Dendrobium candidum, Dendrobium nobile, and Dendrobium fringe were obtained from GenBank. The biological software MEGA 5.0 was used to compare the above sequences to find out all the variation sites of Dendrobium candidum that are different from other species. According to the position of the mutation site, primers of about 20 bases were designed on the upstream and downstream of the ITS2 sequence respectively; the designed primers were analyzed using the biological software Primer Primer 5.0, and finally the appropriate primer pair was determined:

[0039] Upstream primer F: 5'GCTCCGTGCCTTGTCATC3'; and

[0040] Downstream primer R: 5'ATCCACCACTTATTGTACG 3'.

Embodiment 2

[0041] Embodiment 2, utilize PCR primer to detect commercially available Dendrobium officinale sample

[0042] 1. Collect 9 different samples of Dendrobium candidum from the market, take about 30 mg of each sample, add sterilized small steel balls, grind it into powder with a sample mill, and extract DNA by kit method. The kit was purchased from TianGen Company, and the model was Plant Genomic DNA Extraction Kit (spin column type).

[0043] 2. Use ITS universal primers ITS 5F: 5'-GGAAGTAAAAGTCGTAACAAGG-3' and ITS 4R: 5'-TCCTCCGCTTATTGATATGC-3' to perform PCR to detect the quality of each extracted DNA sample.

[0044] PCR reaction system:

[0045]

[0046] The PCR reaction conditions were: 94°C for 5 min; 94°C for 30 sec, 42°C for 1 min, 72°C for 1.22 min, a total of 40 cycles; 72°C for 7 min.

[0047] Set up a blank control and use an equal amount of ddH for the DNA template 2 O instead, prove that the quality of the DNA meets the detection requirements, and determine t...

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Abstract

The invention belongs to the technical field of molecular biology, and discloses an ITS sequence, a specific primer pair, an identification kit, and an identification method used for identification ofdendrobium officinale and high imitation fake products. The ITS sequence possesses a nucleotide sequence represented by SEQ ID NO:1, and can be used for identification of dendrobium officinale and high imitation fake products. The kit used for identification of dendrobium officinale and high imitation fake products is also provided; establishment of a related PCR detection system is carried out,molecular identification basis is provided for dendrobium officinale medicine material identification, rapid accurate detection and distinguish of dendrobium officinale from dendrobium officinale medicine materials with relatively high similarity are realized. Operation is simple and convenient; the specificity is high; the sensitivity is high; detection on dendrobium officinale medicine materialsis realized; and the identification method is suitable for wide popularization and applications.

Description

technical field [0001] The invention belongs to the field of molecular biology, and specifically relates to molecular biology detection of Dendrobium officinale, in particular to an ITS sequence, a pair of specific primers, an identification kit and an identification method for identifying Dendrobium officinale and highly similar counterfeit products. Background technique [0002] Dendrobium is a famous and precious Chinese medicinal material in my country. It was first recorded in "Shen Nong's Materia Medica", listed as the top grade, sweet in nature, slightly cold, and returns to the stomach and kidney meridian. Dendrobium and Dendrobium candidum recorded in the 2015 edition of the "Chinese Pharmacopoeia" are two different varieties of medicinal materials. Wherein, Dendrobium nobile Lindl., Dendrobium chrysotoxum Lindl., Dendrobium fimbriatum Hook. and the fresh or dry stems of the cultivars of Dendrobium nobile Lindl. and similar species of the same genus. Another Dendro...

Claims

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Application Information

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IPC IPC(8): C12Q1/6895C12N15/11
CPCC12Q1/6895
Inventor 余意娄勇军翟旭峰肖小春
Owner INFINITUS (CHINA) CO LTD
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