Method for extracting mitochondria and mitochondrial protein from cotton

A mitochondrial and protein technology, applied in the biological field, can solve problems such as difficult protein extraction

Active Publication Date: 2017-10-20
CHINA AGRI UNIV
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  • Abstract
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Problems solved by technology

Compared with other crops, the extraction of cotton mitochondrial DNA, RNA and protein is mo

Method used

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  • Method for extracting mitochondria and mitochondrial protein from cotton
  • Method for extracting mitochondria and mitochondrial protein from cotton
  • Method for extracting mitochondria and mitochondrial protein from cotton

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0080] Embodiment 1, a kind of method of cotton mitochondria and protein thereof

[0081] 1. Isolation of Cotton Mitochondria

[0082] 1. Mix 100g Xu 244 (Sumian 20) etiolated seedlings with 1000ml Buffer A solution, homogenate, filter with 4-6 layers of gauze, collect the filtrate; centrifuge the filtrate at 1000×g for 10min at low speed, discard the precipitate , take the supernatant; centrifuge the supernatant at 12000×g for 45 min at high speed, discard the supernatant, and collect the precipitate.

[0083] The solvent of above-mentioned Buffer A solution is water, and solute and its concentration in Buffer A solution are: sucrose 300mM, EDTA-Na 2 7.5mM, KCl 10mM, KH 2 PO 4 20mM, BSA 1g / L, dithiothreitol 1.54g / L, β-mercaptoethanol 1ml / L; pH value is 7.5, in which BSA, dithiothreitol and β-mercaptoethanol are added after high temperature and high pressure sterilization.

[0084] 2. Add 40ml Buffer B solution to the precipitate obtained in step 1, brush the precipitate...

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Abstract

The invention discloses a method for extracting mitochondria and mitochondrial protein from cotton. The method has minimum mechanical damage to mitochondria, nucleus components and plasmid components except mitochondria as well as phenolic and polysaccharide substances can be removed effectively, the contamination rate is reduced, the purity and the quality of mitochondrial protein are improved, experimental technique requirements of cotton genome research, mitochondrial proteome research, metabolism regulation network research and the like are met, and related research work fields are broadened.

Description

technical field [0001] The invention belongs to the field of biotechnology, and in particular relates to a method for extracting cotton mitochondria and protein thereof. Background technique [0002] The extraction of mitochondrial protein is mainly divided into two steps: mitochondrial separation and protein extraction. At present, the methods for the separation and extraction of mitochondria in higher plants mainly include differential centrifugation and density gradient centrifugation. Although the differential centrifugation method is simple to operate and is the most widely used method at present, the extracted mitochondria have low purity and are easily contaminated by nuclear impurities, which causes great difficulties for subsequent research work; density gradient centrifugation method, the purity of mitochondria obtained High, but the operation is complicated and time-consuming. Long-term high-speed centrifugation has high requirements for the centrifuge, a large d...

Claims

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Application Information

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IPC IPC(8): C07K1/14
CPCC07K1/145
Inventor 华金平陈志文聂虎帅王梅燕李双双裴海丽
Owner CHINA AGRI UNIV
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