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Extraction method of genomic DNA of plant leaf of traditional Chinese medicine

A technology of plant leaves and extraction method, which is applied in the field of plant genome DNA separation and extraction, and can solve problems such as inability to apply extraction well

Active Publication Date: 2017-05-31
深圳灵天欣元营养科学有限公司
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Problems solved by technology

Therefore, the genome extraction method of one TCM plant is often not well applied to the extraction of genomic DNA of another TCM

Method used

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  • Extraction method of genomic DNA of plant leaf of traditional Chinese medicine

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Embodiment 1

[0046] 1. Extraction method

[0047] Select 0.3 grams each of breviscapine leaves stored at -70°C for one year and fresh;

[0048] 1) Put 0.3 grams of samples in the ice-cold cell nucleus extraction buffer, and use a sharp scalpel to chop the tissue into a very small homogenate mixture;

[0049] 2) Filter the homogenate mixture into a centrifuge tube with a nylon membrane with a pore size of 18-50 μm;

[0050] 3) Centrifuge at 6000-15000rpm for 10-100 seconds at high speed, remove the supernatant, the precipitate is the nucleus, add ice-cold nucleus extraction buffer again to resuspend the precipitate.

[0051] 4) Repeat step 3), 1-5 times;

[0052] 5) Resuspend the cleaned cell nuclei in 3mL cell nucleus lysate, shake until uniform, add 25 μL 20mg / mL proteinase K, shake until uniform, add 300μL 20% SDS, shake well, until viscous, 37°C Digest overnight;

[0053] 6) Add 6 mL of saturated phenol into a 15 mL centrifuge tube, add the overnight digestion solution into the cent...

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Abstract

The invention discloses an extraction method of a genomic DNA of a plant leaf of a traditional Chinese medicine and relates to a separation and extraction technology of genomic DNAs of plants in molecular biology experiments. The method mainly comprises the following steps of: pre-treating a tissue sample; crushing cells; eluting impurities; separating out DNAs; and removing ions. The method disclosed by the invention has the advantages and positive effects that (1) cell nucleuses are released from cell walls by means of a cutting method, then secondary metabolites are eluted to obtain pure cell nucleuses, and finally, the genomic DNA is extracted from the cell nucleuses; (2) liquid nitrogen cannot be used for extracting the cell nucleuses by means of the cutting method; (3) impurities in the cytoplasm are repeatedly eluted, so that pure cell nucleuses can be obtained effectively; (4) samples can be obtained from samples which are stored at a low temperature for a long time and fresh leaves; and (5) the method is suitable for extracting DNAs of all plant leaves of traditional Chinese medicines. The extraction method is suitable for extracting the genomic DNA of the plant leaf of the traditional Chinese medicine.

Description

technical field [0001] The invention relates to a technology for separating and extracting plant genome DNA in molecular biology experiments, in particular to a method for extracting genome DNA from leaves of traditional Chinese medicinal plants. Background technique [0002] Genomic DNA extraction is one of the important links in molecular biology experiments. The quality and yield of genomic DNA directly affect the conduct of subsequent DNA-related experiments in molecular biology experiments. Chinese medicinal plants contain a large number of secondary metabolites, especially starch, polyphenols and polysaccharides, which seriously affect the quality of extracted DNA. The types of secondary metabolites in this kind of plants are complex, and the heterogeneity of secondary metabolites among different species and even in different seasons of the same species is very high. Therefore, the genome extraction method of one TCM plant is often not well applied to the extraction ...

Claims

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Application Information

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IPC IPC(8): C12N15/10
CPCC12N15/1003
Inventor 胡军军马璐
Owner 深圳灵天欣元营养科学有限公司
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