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Plant genome DNA extraction method and application thereof

A plant genome and extraction method technology, applied in the field of plant genome DNA extraction, can solve the problems of incomplete removal of impurities such as polysaccharides and polyphenols, threats to the safety of experimenters, and low DNA purity, achieving excellent cost and time advantages, Meet the requirements of genome sequencing and the effect of simple operation steps

Active Publication Date: 2019-12-20
BIOMARKER TECH
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  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Because the phenol and chloroform extraction reagents used in the traditional CTAB method are toxic, the safety of the experimenters is greatly threatened
However, the alternative methods of the CTAB method reported in the prior art may have the problems of high cost and unsuitability for processing a large number of samples (such as magnetic bead purification, column purification method), or incomplete removal of impurities such as polysaccharides and polyphenols, resulting in DNA purity. Lower problems (eg SDS fragmentation method)

Method used

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  • Plant genome DNA extraction method and application thereof
  • Plant genome DNA extraction method and application thereof
  • Plant genome DNA extraction method and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0043] Embodiment 1 A kind of extraction method of plant genomic DNA

[0044] The present embodiment provides a method for extracting plant genomic DNA, specifically as follows (flow chart as follows figure 1 shown):

[0045] 1. After thoroughly cooling the mortar and grinding rod in liquid nitrogen, put the plant tissue sample into the mortar, immediately inject liquid nitrogen into the mortar, and fully grind the tissue into powder with the mortar rod (guarantee the sample It cannot be melted and needs to be kept at a low temperature); after fully grinding, put 100mg of tissue powder into 2.0ml imported centrifuge tubes that have been pre-cooled and marked with the name of the sample immediately with a liquid nitrogen pre-cooled spoon, and tightly cover the tube cap Then put it in liquid nitrogen and save it for later use (the liquid nitrogen in the centrifuge tube must be fully volatilized before loading to prevent the tube from blowing up).

[0046] 2. Add 1 mL of 65°C p...

experiment example 1

[0055] EXTRACTION AND QUALITY ANALYSIS OF EXPERIMENTAL EXAMPLE 1 Wheat Genomic DNA

[0056] Utilize the method of embodiment 1 and comparative example 1 to extract the genomic DNA of wheat leaf respectively, and utilize Nanodrop, Qubit and agarose gel electrophoresis to utilize the method that embodiment 1 and comparative example 1 provides to extract the genomic DNA of wheat leaf that obtains concentration , purity and integrity analysis.

[0057] The detection results of the concentration and purity of wheat leaf genomic DNA are as shown in Table 1, and the results show that the extraction method of embodiment 1 and the extraction method of comparative example 1 extract the wheat leaf genomic DNA obtained in concentration and purity. The method of Example 1 of the present invention extracts wheat genomic DNA with high yield and high purity, which can meet the requirements of NGS sequencing for the purity and total amount of DNA samples.

[0058] Table 1 Quality detection of...

experiment example 2

[0061] Experimental example 2 Extraction and quality analysis of genomic DNA of other crops

[0062] Using the method of Example 1 to extract genomic DNA from the leaves of seven plants, namely rice, cotton, sorghum, soybean, peanut, millet and corn, and use Nanodrop, Qubit and agarose gel electrophoresis to extract the obtained DNA using the method of Example 1 Genomic DNA from leaves of rice, cotton, sorghum, soybean, peanut, millet and maize was analyzed for concentration, purity and integrity.

[0063] The detection results of the concentration and purity of rice, cotton, sorghum, soybean, peanut, millet and corn genomic DNA are shown in Table 2, and the results show that the method of Example 1 of the present invention is used to extract rice, cotton, sorghum, soybean, peanut, Genomic DNA from millet and maize leaves has a high yield and good purity, which can meet the requirements of NGS sequencing for the purity and total amount of DNA samples.

[0064] Table 2 Quality...

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Abstract

The present invention relates to the technical field of molecular biology and particularly relates to a plant genome DNA extraction method and an application thereof. The provided plant genome DNA extraction method comprises the following steps: after utilizing CTAB to lyse plant cells, adding SDS and EDTA solutions for extraction, and performing precipitation to obtain genome DNA. The method usesa CTAB lysis reagent to lyse plant tissue cells, the SDS and EDTA are cooperated to be used as extraction reagents to precipitate and remove impurities such as proteins. The provided plant genome DNAextraction method can obtain plant genome DNA with high concentration, high purity and high integrity, can meet requirements of genome sequencing, at the same time avoids uses of phenol / chloroform toxic chemical reagents, also has advantages of low cost, simple operation steps, short extraction time, and is suitable for genome DNA extraction of a large number of plant sample genome sequencing.

Description

technical field [0001] The invention relates to the technical field of molecular biology, in particular to a plant genome DNA extraction method and application thereof. Background technique [0002] With the development of genome sequencing technology, the research on the sequence, structure and function of plant genome is developing rapidly, and obtaining high-purity, high-content and high-integrity plant genomic DNA is the primary prerequisite for the application of genome sequencing technology. Since plant tissues contain a large amount of polysaccharides, polyphenols and other secondary metabolites with complex structures, the extraction of plant genomic DNA is more difficult than the extraction of genomes from microorganisms, animal tissues or blood samples. In addition, the construction of genome sequencing library has higher requirements for the integrity, purity and concentration of genomic DNA than ordinary experiments such as templates used for detection amplificat...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/10C12Q1/6806C40B50/06
CPCC12N15/1003C12Q1/6806C40B50/06C12Q2523/32
Inventor 郑洪坤毕经德骆晨朱艳荣王瑞
Owner BIOMARKER TECH