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Universal kit and method for extracting DNA from trace plant materials

A plant material and kit technology, which can be used in DNA preparation, recombinant DNA technology, and microbial assay/inspection, etc., and can solve problems such as time-consuming and labor-intensive

Inactive Publication Date: 2017-10-20
北京爱普拜生物技术有限公司
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Usually, the total amount of DNA extracted from a 100mg plant sample is 10-15μg, and the DNA yield rate is 0.01-0.15%. More than one extraction reaction is often required to meet the needs of whole-genome sequencing, which is time-consuming and labor-intensive.

Method used

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  • Universal kit and method for extracting DNA from trace plant materials
  • Universal kit and method for extracting DNA from trace plant materials
  • Universal kit and method for extracting DNA from trace plant materials

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0078] The DNA extraction kit of embodiment 1 whole genome sequencing of plants is to the extraction result of the plant material of different weights

[0079] 1. Experimental method

[0080] Take 0.0001, 0.0005, 0.001, 0.005, 0.01, 0.05, 0.1, 0.5, and 1 gram of fresh soybean leaf tissue respectively, and add 600 μL of AP1 Buffer and 6 μL RNaseA (10 mg / mL), machine-polished samples in a grinding tube; 0.5 and 1 g of leaf samples were fully ground into a fine powder by adding liquid nitrogen in a mortar and transferred the fine powder to a 1.5 mL centrifuge tube, Add 600 μL AP1 buffer and 6 μL RNaseA (10 mg / mL), place at 65°C for 10 minutes, invert the centrifuge tube 2 to 3 times during the standing process, and mix the sample;

[0081] Add 200μL AP2 buffer, mix well, centrifuge at 13,000rpm for 5 minutes, carefully pipette the supernatant into a new 1.5mL centrifuge tube;

[0082] Add 1.5 times the volume of supernatant AP3 buffer, and immediately mix thoroughly by pipettin...

Embodiment 2

[0097] The DNA extraction kit of embodiment 2 whole genome sequencing of plants is to the extraction result of different kinds of plant materials

[0098] 1. Experimental method

[0099] Take 5 mg each of barley leaves, corn leaves, rice leaves (Japanese nitrile), sorghum leaves, soybean leaves, tomato leaves, dandelion leaves, wild chrysanthemum leaves, pothos leaves, gladiolus leaves, and pine needles, and add 600 μL of AP1 buffer and 6 μL of RNaseA (10mg / mL), use a proofer to crush the sample, place it at 65°C for 10 minutes, and mix the sample by inverting the centrifuge tube 2 to 3 times during the placement process;

[0100] Add 200 μL AP2 buffer, mix thoroughly, centrifuge at 13,000 rpm for 5-10 minutes, and carefully pipette the supernatant into a new 1.5mL centrifuge tube;

[0101] Add 1.5 times the volume of supernatant AP3 buffer, and immediately mix thoroughly by pipetting. If flocculent precipitation occurs at this time, it will not affect DNA extraction;

[010...

Embodiment 3

[0115] Example 3 Extraction Results of Different Parts of Plant Material by the DNA Extraction Kit for Plant Whole Genome Sequencing

[0116] 1. Experimental method

[0117] Take 5 mg of different parts of the root, stem, flower, fruit, and seed of the plant, add 600 μL of AP1 buffer solution and 6 μL of RNaseA (10 mg / mL), break the sample with a proofer, place it at 65 ° C for 10 minutes, and centrifuge it upside down during the placement process tube 2 to 3 times, mix the sample;

[0118] Add 200 μL AP2 buffer, mix thoroughly, centrifuge at 13,000 rpm for 5-10 minutes, and carefully pipette the supernatant into a new 1.5mL centrifuge tube;

[0119] Add 1.5 times the volume of supernatant AP3 buffer, and immediately mix thoroughly by pipetting. If flocculent precipitation occurs at this time, it will not affect DNA extraction;

[0120] Put the mixture (solution and possible flocculent precipitate) obtained in the previous step into an adsorption column DB (the adsorption co...

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Abstract

The invention belongs to the field of molecular biology and mainly relates to a universal kit and method for extracting DNA from trace plant materials. The yield of DNA extracted once through the kit and the method can completely meet the requirement of direct whole genome sequencing. The kit and the method mainly comprise a thin silicon matrix adsorption column capable of adsorbing genome DNA to the maximum extent and a unique extraction buffering liquid system which is matched with the silicone matrix adsorption column and can extract various plant tissues and DNA in different parts of plants to the maximum extent. The kit and the method have the advantages of rapidness, convenience, no need of phenol-chloroform extraction, avoidance of organic solvent pollution and the like, and pigments, grease and other impurities in the DNA extraction process can be simply and conveniently removed to the maximum extent due to the thin adsorption column. Valuable plant samples are saved, and pretreatment steps and time of whole genome sequencing are greatly reduced and shortened.

Description

technical field [0001] The patent of the present invention relates to a general-purpose kit and method for extracting DNA from trace plant materials. This method can extract a large amount of genomic DNA from common plant materials, especially trace plant materials, to meet the needs of whole-genome sequencing of precious small samples. requirements. Background technique [0002] Genome is the overall composition of all genes in a species, which carries the genetic material and genetic information of this species. To uncover the mysteries of life, it is necessary to study the existence of genes, the structure and function of genes, and the interrelationships between genes at the overall level. Whole genome sequencing is to sequence all the genes in the genome of an organism, determine the base sequence of its DNA, and detect all the genetic information in the individual genome. Through whole-genome sequencing of the genome, the genome sequence map of the species can be obt...

Claims

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Application Information

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IPC IPC(8): C12N15/10
CPCC12N15/101C12Q2523/308
Inventor 于祥春冯晓燕林挺王文利龚建
Owner 北京爱普拜生物技术有限公司
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