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Wu-he dipsacus asper ISSR-PCR molecule marking method

A technique of molecular markers and discontinuities, applied in the field of molecular biology, can solve the problems of little research on genetic diversity and achieve the effects of easy and rapid analysis and detection, good polymorphism characteristics, and clear and stable bands

Inactive Publication Date: 2017-02-22
湖北省农业科学院中药材研究所
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Problems solved by technology

[0003] Current research mainly focuses on historical research, development prospects, karyotype analysis, component analysis, cultivation characteristics, etc., while there are few basic researches on physiology, biochemistry and genetic diversity.

Method used

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  • Wu-he dipsacus asper ISSR-PCR molecule marking method

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Embodiment 1

[0026] A method for ISSR-PCR molecular marker of Dipsacus sativa of the present invention comprises the following steps:

[0027] (1) Sampling and Genomic DNA Extraction

[0028] Wash the ground mortar with water and dry it, add 5ml of absolute ethanol and ignite it, add liquid nitrogen to pre-cool after the fire is extinguished, select 0.5g of seedlings that have germinated for 15-20 days in the room, remove the seed shell and put Put it into a pre-cooled mortar, add an appropriate amount of liquid nitrogen, cover the sample with liquid nitrogen for 2-3 cm, grind for 3-5 minutes, and then use the kit method to extract the genomic DNA of Dipsacus sativa.

[0029] (2) Uniform design scheme for the first round of ISSR-PCR reaction system

[0030] Uniform optimization design screens the most suitable amount of each component of the ISSR reaction system, using U 12 (6 2 ×4) Mixed uniform design table, the factor levels of the PCR amplification system are shown in Table 2. Usin...

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Abstract

The invention relates to a wu-he dipsacus asper ISSR-PCR molecule marking method. The method is characterized by comprising the following steps: 1, extracting the genome of the wu-he dipsacus asper, 2, conducting 1% agarose electrophoresis for 20 minutes, testing the integrity of the genome, at the same time using the DNA bar code ITS2 primer to conduct the PCR amplification of the extracted wu-he dipsacus asper DNA, verifying the purity of the extracted DNA, 3, based on the initial screened primer conducting the ISSR-PCR. Compared with the prior art, the reaction system created by the method has a simple optimization method. The amplified bands is both clear and stable with good polymorphism. The method compensates the shortcomings in the study of the genetic diversity of the wu-he dipsacus asper. The method can be used in the analysis of the genetic relationship and the verification of authenticity of the teasle, thus having good application value.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to an ISSR-PCR molecular marker method for Dipsacus wuheensis. Background technique [0002] Dipsacus is a commonly used traditional Chinese medicine, which is the dried root of Dipsacus asper Wall. It has the effects of nourishing liver and kidney, strengthening muscles and bones, stopping bleeding and preventing miscarriage; while Dipsacus, which is produced in Hefeng County, Enshi Prefecture, Hubei Province, and Wufeng County, Yichang City, because of its strong roots, uniform thickness, and soft texture, is excellent. Known as "Five Crane Dipsacus", it is one of the important authentic medicinal materials in Enshi Prefecture. Existing studies have shown that the quality of the medicinal materials of Dipsacus chuanxiong not only varies with different places of origin, but also has obvious differences in biological characteristics among different populations, suggesting that Dips...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/68
CPCC12Q1/6895C12Q2600/156
Inventor 卢超艾伦强张美德何银生刘海华
Owner 湖北省农业科学院中药材研究所
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