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Technology for extracting genomic deoxyribonucleic acid (DNA) from recalcitrant plant

A technology of plant genome and technology, which is applied in the field of "recalcitrant" plant genome DNA extraction technology, can solve the problem of less use of mature leaves, and achieve the effect of solving stickiness and avoiding browning

Inactive Publication Date: 2012-12-05
刘军
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Problems solved by technology

Because young leaves have less secondary metabolites such as polysaccharides and polyphenols, and genomic DNA extraction is easier, so the plant materials used in previous studies were mostly young leaves, and less mature leaves and silica gel drying leaves were used.

Method used

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Embodiment Construction

[0009] Below in conjunction with specific embodiment the present invention is described in further detail:

[0010] 1 Solution preparation

[0011] Elution buffer: 100mM Tris-HCl, pH 8.0; 1.4M NaCl; 20mM EDTA, pH=8.0; 2% β-mercaptoethanol (V / V); 2% PVP (W / V) (polyvinylpyrrole Alkanone K-40); β-mercaptoethanol and PVP are added before use;

[0012] 2×CTAB extraction buffer: add 2% CTAB (W / V) to the elution buffer;

[0013] For other reagents, refer to conventional preparation methods.

[0014] 2 Extraction process

[0015] 2.1 Put about 0.1g of shredded fresh leaves or about 0.05g of dried silica gel leaves into an agate mortar, add a small amount of quartz sand, and then pour a little liquid nitrogen, and grind it vigorously when the liquid nitrogen is about to evaporate, and put The ground leaves were added to the centrifuge tube previously filled with 1mL elution buffer, mixed upside down and ice-bathed for 5min;

[0016] 2.2 Centrifuge the centrifuge tube in a high-spe...

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Abstract

The invention provides a technology for extracting genomic deoxyribonucleic acid (DNA) from a recalcitrant plant, and belongs to the technical field of biology. The technology is characterized in that secondary metabolism substances such as polysaccharide and polyphenol can be successfully separated from the recalcitrant plant by utilizing eluent, and the problem of ropiness of a centrifugal supernatant is solved; phenol, chloroform and isoamyl alcohol are utilized to conduct the extraction, then the chloroform and the isoamyl alcohol are utilized to conduct the extraction, and protein, the phenol, and the like can be completely extracted; and polyphenol substances can be prevented from being oxidized into quinines and the browning is avoided, so that high-quality genomic DNA of the of the recalcitrant plant is successfully extracted. The technology is mainly applied to the extraction of genomic DNA of plant materials of which cells contain plenty of secondary metabolite such as the polysaccharide, the polyphenol, tanned matters, cinnamaldehyde, cinnamic acid, and the like.

Description

1. Technical field [0001] The patent of the present invention belongs to the field of biotechnology, and relates to a "recalcitrant" plant genome DNA extraction technology, which can significantly improve the quality of genome DNA, and is mainly used in cells containing a large amount of polysaccharides, polyphenols, keratin and cinnamon aldehyde, Genomic DNA extraction from plant material of secondary metabolites such as cinnamic acid. 2. Background technology [0002] Because the cells of "recalcitrant" plants such as Machilus thunbergii Sieb.et Zucc., Machilus leptophylla Hand.-Mazz. and Longan (Dimocarpus longan Lour.) contain a large amount of polysaccharides, polyphenols, and And secondary metabolites such as cinnamon aldehyde and cinnamon acid can not only interfere with the DNA extraction process, but also make the extracted DNA solution viscous, and the sample is brown, so it cannot be used as a PCR template for molecular biological analysis and research. Molecular ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
Inventor 刘军
Owner 刘军
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