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Method for extracting rice genome deoxyribonucleic acid (DNA)

An extraction method and genome technology, applied in the field of simple extraction of high-purity, large-segment rice genomic DNA, to achieve the effect of reducing the number, reducing the number of centrifuges and rotating speeds, and simplifying the operation steps

Inactive Publication Date: 2014-02-05
HENAN NORMAL UNIV
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  • Claims
  • Application Information

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Problems solved by technology

At present, there is no report on the simple preparation method of high-purity and large-segment rice genomic DNA with a length of at least 100 kb that meets the requirements for the preparation of genomic libraries.

Method used

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  • Method for extracting rice genome deoxyribonucleic acid (DNA)

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Experimental program
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Embodiment 1

[0020] (1) Take 1 gram of fresh rice yellow flower seedling material and grind it with liquid nitrogen. Before thawing, transfer the powder to 2ml of CTAB buffer solution (in a water bath) that has been preheated to 65°C. Gently rotate the centrifuge tube and mix gently. 65 ℃ water bath for 2h. Among them, CTAB buffer: 100mM Tris-HCl (pH8.0), 20mM EDTA, 1.4M NaCl, 2% CTAB, add 0.2% β-mercaptoethanol before use;

[0021] (2) Take the cleavage mixture out of the water bath, cool to room temperature, add an equal volume of chloroform / isoamyl alcohol (the volume ratio of the two is 24:1), and gently invert to form an emulsion.

[0022] (3) Centrifuge at room temperature at 5000rpm for 10min, transfer the upper aqueous phase to a new tube, add 1 / 100 volume of RNase A (10mg / ml), gently invert and mix, and incubate at 37°C for 30min.

[0023] (4) Add 0.7 times the volume of isopropanol and mix carefully. Once white filaments appear, immediately hook them out with a glass hook.

[0...

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Abstract

The invention discloses a method for extracting rice genome deoxyribonucleic acid (DNA). The method comprises the steps of splitting, extracting, carrying out centrifugal separation and enzymolysis, precipitating, etc. After the method is adopted, the large-fragment rice genome DNA can be simply and effectively extracted; compared with the prior art, the method is simplified in the operation steps, reduced in the centrifugation times and rotating speed and reduced in the number of the needed reagents; the quality of the DNA completely meets the common subsequent experiment requirements such as genomic library building, genomic sequence amplification, Southern blot, tail-polymerase chain reaction (PCR), etc.

Description

technical field [0001] The invention belongs to the field of biology, and in particular relates to a simple extraction method of high-purity and large-segment rice genomic DNA. Background technique [0002] Rice is not only a major food crop, but also an important model organism. With the development of rice genome research, molecular biology techniques have been widely used in rice research. In rice molecular biology research, extracting and preparing its DNA is the basis and premise of many research work, such as: constructing genomic library, Southern hybridization, RFLP, PCR to isolate genes and regulatory regions, etc. In these genomic DNA-based operations, the quality requirements for extracted DNA include not only high yield, high purity, good integrity, and as little fragmentation and degradation as possible, but also the exclusion of other macromolecules, such as proteins, polysaccharides, etc. Contamination . Since some plant tissue materials are rich in polysac...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10
Inventor 梁卫红李佳佳楼晨林群婷杨丹丹王俊杰郝雨帆
Owner HENAN NORMAL UNIV
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