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Kit for extracting total DNA from citrus leaf midribs on basis of magnetic bead method and extraction method thereof

A magnetic bead method and kit technology, applied in the field of analytical biology, can solve the problems that have not been developed, and achieve the effects of low cost, high concentration and purity, and high detection sensitivity

Active Publication Date: 2019-04-05
ZHONGKAI UNIV OF AGRI & ENG
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, the market mainly sells DNA extraction kits with magnetic beads for plants or animals, and a DNA extraction kit with magnetic beads for citrus Huanglongbing has not been developed yet.

Method used

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  • Kit for extracting total DNA from citrus leaf midribs on basis of magnetic bead method and extraction method thereof
  • Kit for extracting total DNA from citrus leaf midribs on basis of magnetic bead method and extraction method thereof
  • Kit for extracting total DNA from citrus leaf midribs on basis of magnetic bead method and extraction method thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0042] A kit for extracting total DNA from the midrib of citrus leaves based on the magnetic bead method, comprising a lysate, a binding solution, a washing solution 1, a washing solution 2, an eluent and a magnetic bead suspension;

[0043] The lysate consists of: 100mmol / L pH=8.0Tris-HCl, 25mmol EDTA, 0.14mol / LNaCl, 3%m / v CTAB, 1%m / v potassium hydrogensulfate, 1%m / v ascorbic acid, 2 % m / v PVP-40T, 0.1% v / v β-mercaptoethanol, solvent is ddH 2 O, made into 100 mL; among them, potassium bisulfate, PVP-40T, and β-mercaptoethanol were added before use.

[0044] The preparation of the lysate is as follows: the above-mentioned Tris-HCl, EDTA, NaCl, CTAB, potassium bisulfate, ascorbic acid, PVP-40T, β-mercaptoethanol are mixed, dissolved in ddH 2 O, made into 100mL, that is, potassium bisulfate, PVP-40T, and β-mercaptoethanol were added before use.

[0045] The composition of the binding solution is: 750mmol / L NaCl, 50mmol / L pH=7.0MOPS, 0.15% v / v Triton X-100, 15% v / v isopropanol,...

Embodiment 2

[0056] A kit for extracting total DNA from the midrib of citrus leaves based on the magnetic bead method, comprising a lysate, a binding solution, a washing solution 1, a washing solution 2, an eluent and a magnetic bead suspension;

[0057] The lysate consists of: 80mmol / L pH=7.5Tris-HCl, 15mmol EDTA, 0.1mol / LNaCl, 2%m / v CTAB, 0.5%m / v potassium hydrogensulfate, 1.5%m / v ascorbic acid, 3.0 % m / v PVP-40T, 0.2% v / v β-mercaptoethanol, solvent is ddH 2 O, made into 100 mL; among them, potassium bisulfate, PVP-40T, and β-mercaptoethanol were added before use. The preparation of the lysate refers to Example 1.

[0058] The composition of the binding solution is: 600mmol / L NaCl, 60mmol / L pH=6.0MOPS, 0.15% v / v TritonX-100, 10% v / v isopropanol, and the solvent is ddH 2 O, made into 100 mL; where, isopropanol was added before use. The preparation of the binding solution refers to Example 1.

[0059] The composition of the washing solution 1 is: 6 mol / L guanidine hydrochloride, 10 mmo...

Embodiment 3

[0064] A kit for extracting total DNA from the midrib of citrus leaves based on the magnetic bead method, comprising a lysate, a binding solution, a washing solution 1, a washing solution 2, an eluent and a magnetic bead suspension;

[0065] The composition of the lysate is: 120mmol / L pH=8.5Tris-HCl, 30mmol EDTA, 0.2mol / LNaCl, 4%m / v CTAB, 1%m / v potassium hydrogensulfate, 0.5%m / v ascorbic acid, 1.0 % m / v PVP-40T, 0.1% v / v β-mercaptoethanol, solvent is ddH 2 O, made into 100 mL; among them, potassium bisulfate, PVP-40T, and β-mercaptoethanol were added before use. The preparation of the lysate refers to Example 1.

[0066] The composition of the binding solution is: 800mmol / LNaCl, 40mmol / L pH=7.0MOPS, 0.15% v / v TritonX-100, 20% v / v isopropanol, and the solvent is ddH 2 O, made into 100 mL; where, isopropanol was added before use. The preparation of the binding solution refers to Example 1.

[0067] The composition of the washing solution 1 is: 4mol / L guanidine hydrochloride,...

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Abstract

The invention discloses a kit for extracting total DNA from citrus leaf midribs on the basis of a magnetic bead method and an extraction method thereof. The kit comprises cracking liquid, combining liquid, washing liquid 1, washing liquid 2, eluate and magnetic bead suspension liquid. When the DNA extraction kit provided by the invention is used for extracting the total DNA from citrus leaves, theoverall levels of the concentration and the purity of the extracted DNA are superior to those of a Sangon Dzup (plant) genome DNA fast extraction kit and a Tiangen plant DNA extraction kit sold in the market. Aiming at citrus yellow shoot detection, the invention provides the DNA extraction magnetic bead method kit and the extraction method of the kit. The kit and the extraction method are applicable to all citrus varieties; board spectrum performance is realized.

Description

technical field [0001] The invention relates to the technical field of analytical biology, in particular to a kit for extracting total DNA from midribs of citrus leaves based on a magnetic bead method and an extraction method thereof. Background technique [0002] Citrus huanglongbing is a devastating disease of citrus, which has caused huge losses to the world's citrus industry. So far, the prevention and control of citrus Huanglongbing at home and abroad mainly relies on the main measures such as eliminating vectors, strengthening quarantine, cultivating disease-free seedlings, and digging up diseased trees. The prevention and control measures of citrus huanglongbing need the help of precise diagnosis technology for citrus huanglongbing. PCR detection of huanglongbing pathogen "Candidatus Liberibacter spp." is currently a reliable method for the diagnosis of citrus huanglongbing, which has the advantages of rapidity, specificity, efficiency and sensitivity. [0003] At pr...

Claims

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Application Information

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IPC IPC(8): C12N15/10C12Q1/6806
CPCC12N15/1013C12Q1/6806C12Q2563/143C12Q2563/149
Inventor 吴仲真林进添宾淑英康聪蒲新华吴泳贤
Owner ZHONGKAI UNIV OF AGRI & ENG
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