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Veratrum ISSR-PCR molecular marking method

A technology of molecular markers and veratrum, applied in the field of biology, can solve the problems of unreported research on molecular genetics and germplasm resources of veratrum, and achieve a stable and reliable reaction system

Pending Publication Date: 2019-09-03
INNER MONGOLIA MEDICAL UNIV
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AI Technical Summary

Problems solved by technology

[0005] At present, the researches on veratrum mainly focus on the chemical composition, quality analysis and pharmacological effects, and there are no reports on the molecular genetics and germplasm resources of veratrum.

Method used

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  • Veratrum ISSR-PCR molecular marking method
  • Veratrum ISSR-PCR molecular marking method

Examples

Experimental program
Comparison scheme
Effect test

Embodiment

[0026] 1 Genomic DNA extraction and detection

[0027] Use Tiangen spin column type DNA extraction kit to extract Veratrum genomic DNA, 0.8% agarose gel electrophoresis, and detect DNA quality according to the imaging results; use UV spectrophotometer to measure DNA concentration and purity, after appropriate dilution, place in- Store in a refrigerator at 20°C for later use.

[0028] 2 Optimization of Veratrum ISSR-PCR reaction system

[0029] 2.1 Orthogonal experiment to optimize the PCR system

[0030] In this experiment, L16(4 5 ) to design an orthogonal experiment to influence the template DNA concentration, primer concentration, Taq DNA polymerase concentration, Mg 2+ Concentration and dNTP concentration were carried out in an orthogonal experiment with 5 factors and 4 levels (Table 1), with 16 treatments in total, and 3 repetitions for each treatment. In this experiment, UBC856 was selected as the ISSR-PCR amplification primer, and each reaction system was sterilized...

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Abstract

The invention discloses a veratrum ISSR-PCR molecular marking method; with veratrum genomic DNA as a template and UBC856 as a primer and with adopting of 20 [mu]L of an ISSR-PCR reaction system, the amplification procedures comprise the steps: carrying out pre-denaturation for 2 min at the temperature of 94 DEG C, carrying out denaturation for 30 s at the temperature of 94 DEG C, annealing for 30s at the temperature of 51.8 DEG C, extending for 30 s at the temperature of 72 DEG C, circulating for 30 times, extending for 2 min at the temperature of 72 DEG C, and preserving at the temperature of 4 DEG C; an ISSR-PCR reaction system comprises 0.5 [mu]mol / L of the primer, 24 ng / 20 [mu]L of the template DNA, 1 U / 20 [mu]L of Taq DNA polymerase, 0.2 mmol / L of dNTP, and 1.0 mmol / L of Mg<2+>. Thereaction system is stable and reliable, is suitable for subsequent genetic diversity analysis and genetic variation process and evolution research of veratrum, and provides reference basis for germplasm resource identification, genetic relationship analysis, genetic map research and the like.

Description

technical field [0001] The invention relates to the field of biology, in particular to a veratrum ISSR-PCR molecular marker method. Background technique [0002] Veratrum ( Veratrum nigrum L.) is a plant of the genus Veratrum of the Liliaceae family, a perennial herb, and is one of the more commonly used Chinese and Mongolian medicine varieties in China. Widely distributed in Inner Mongolia, Heilongjiang, Jilin, Tibet, Qinghai, Gansu, Sichuan and Yunnan, the habitats are mostly forest source and meadow. [0003] The dried roots and rhizomes of veratrum are used in Mongolian medicine, named "Age Xiriga", which is used in the "clearing and purging therapy" characteristic of Mongolian medicine to induce vomiting and clearing diarrhea, and to treat "Sheila" disease, indigestion, and lumps disease etc. According to the traditional theory of Mongolian medicine, many tumors and cancers clinically diagnosed by modern medicine belong to the lump disease defined by Mongolian medic...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/6895C12Q1/6858
CPCC12Q1/6895C12Q1/6858C12Q2600/156C12Q2531/113
Inventor 张屏岳鑫包保全唐雨晨程杰宝音萨础拉吴双鹰晓花赵红梅
Owner INNER MONGOLIA MEDICAL UNIV
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