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Method for effectively extracting plant mitochondria

A mitochondrial and thick mitochondrial technology, applied in plant cells, etc., can solve the problems of chloroplast debris interference, mitochondria prone to adhesion, hindering the study of plant mitochondrial function, etc., and achieve the effect of eliminating the interference of chloroplasts

Active Publication Date: 2020-11-20
PEKING UNIV
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Problems solved by technology

[0003] In the prior art, the method of combining differential centrifugation and density gradient centrifugation is often used to separate the mitochondria of plant tissues. The steps of this method are cumbersome, and the mitochondria are prone to adhesion and interference of chloroplast fragments during the separation and purification process, which hinders the functional research of plant mitochondria.

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  • Method for effectively extracting plant mitochondria
  • Method for effectively extracting plant mitochondria

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[0025] The implementation of the present invention will be described in detail below in conjunction with the drawings and examples.

[0026] Such as figure 1 As shown, a method for effectively extracting plant mitochondria of the present invention, differential centrifugation combined with an inorganic alumina filter membrane, for plant mesophyll cells, to separate and purify mitochondria, the main steps are:

[0027] 1) Cultivation of plant materials: AtDIPS-GFP single-labeled transgenic tobacco or Arabidopsis were cultured in plug trays, in terms of mass, nutrient soil: compressed soil: vermiculite = 1:1:1, temperature 23 + 1°C, 16-20h light, 8-10h dark culture, 8 weeks for tobacco, 4 weeks for Arabidopsis, pay attention to replenish water in time. The growth state of the material is an important factor for the success or failure of plant mitochondrial extraction.

[0028] 2) Preparation of grinding buffer:

[0029] ① Grinding buffer (Buffer A): 300mM sucrose (to maintain...

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Abstract

The invention discloses a method for effectively extracting plant mitochondria. The method is characterized by combining differential centrifugation with an inorganic alumina filter membrane, mainly aims at plant mesophyll cells, efficiently separates and purifies mitochondria, and comprises the following steps: 1) culturing of a plant material; 2), preparation of a grinding buffer solution; 3), sample preparation; 4), syringe needle filter membrane filtration; and 5), fluorescence microscope mitochondrion detection. The method is convenient to operate and stable in effect, and has important significance in plant mitochondrion function exploration.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular to a method for effectively extracting plant mitochondria. Background technique [0002] Mitochondria are the "power processing plants" of cell life activities, and have the characteristics of semi-autonomous inheritance. Many vital processes of cell life activities are related to mitochondria, such as cell differentiation, cell information transmission and cell apoptosis, and have the ability to regulate cell growth and cell cycle. The interference of chloroplasts in plant cells has caused great obstacles to the purification of mitochondria, especially in mesophyll cells. In previous studies, in order to avoid the interference of plastids such as chloroplasts, etiolated seedlings or roots or suspension cells without chloroplasts were mostly selected as experimental materials. Using the mechanism of different densities of mitochondria and chloroplasts, sucrose or percoll gradient densi...

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Application Information

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IPC IPC(8): C12N5/04
CPCC12N5/04
Inventor 于婷乔段光兴袁英张立光张泉苏都莫日根
Owner PEKING UNIV
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