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Method for efficiently enriching plant mitochondria through protoplast

A protoplast, enriching plant technology, applied in plant cells and other directions, can solve the problems of cumbersome extraction process, unfavorable rapid acquisition of mitochondria, etc., and achieve the effect of high purity and elimination of interference.

Inactive Publication Date: 2020-11-27
PEKING UNIV
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  • Abstract
  • Description
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Problems solved by technology

However, such an extraction process is too cumbersome and requires ultra-high-speed centrifugation, which is not conducive to the rapid acquisition of mitochondria

Method used

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  • Method for efficiently enriching plant mitochondria through protoplast
  • Method for efficiently enriching plant mitochondria through protoplast
  • Method for efficiently enriching plant mitochondria through protoplast

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Embodiment Construction

[0028] The implementation of the present invention will be described in detail below in conjunction with the drawings and examples.

[0029] like figure 1 As shown, the present invention efficiently enriches plant mitochondria by protoplasts. After obtaining pure mitochondria, the inorganic alumina filter membrane is utilized in conjunction with differential centrifugation to target plant mesophyll cells (or other plant tissues that are easy to obtain protoplasts, other Plant tissues that can extract protoplasts can refer to this process), and the main steps of isolating and purifying mitochondria are:

[0030] 1) Cultivation of plant materials: Taking the leaves of tobacco (Nicotiana tabacum) as an example, mature and plump seeds of AtDIPS-GFP single-label transgenic tobacco were cultivated in plug trays, and the composition of the cultivation soil was nutrient soil by mass: compressed Soil: vermiculite = 1:1:1, temperature 23+1°C, 16 hours of light, 8 hours of dark cultivat...

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Abstract

The invention relates to a method for efficiently enriching plant mitochondria through protoplast. The technical core of the method is acquisition of the protoplast and filtration of a filter membrane, and mitochondria are efficiently separated and purified mainly aiming at plant mesophyll cells and other tissue components capable of dissociating the protoplast. The method comprises the followingsteps: 1) culturing a plant material; 2) dissociating the protoplast; 3) naturally spalling the protoplast by using osmotic pressure; 4) performing differential centrifugation to obtain crude mitochondria; 5) filtering a heavy suspension buffer of the crude mitochondria with a needle filter made of PES material to obtain pure mitochondria; and 6) detecting the purity of the mitochondria through afluorescence microscope. The method is convenient to operate, long in consumed time, stable in effect and capable of efficiently enriching the mitochondria, and has important significance in exploringthe mitochondria functions of plants.

Description

technical field [0001] The invention belongs to the field of biotechnology, in particular to a method for efficiently enriching plant mitochondria through protoplasts. Background technique [0002] Mitochondria and chloroplasts are the energy centers of cellular activities. Chloroplasts supply energy to biological organisms through photosynthesis. Mitochondria convert the energy stored in organic matter into ATP, the direct energy source for cellular life activities, through oxidative phosphorylation. Both mitochondria and chloroplasts have semi-autonomous inheritance characteristics, and together with the nucleus, they constitute the genetic pool of plants. Mitochondria and chloroplasts are complementary and inseparable organelle components in plant organisms. However, in order to study the structure and function of plant mitochondria, the existence of chloroplasts also sets obstacles for the study of mitochondria. In previous studies, in order to avoid the interference o...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/04
CPCC12N5/04
Inventor 于婷乔段光兴袁英张泉苏都莫日根
Owner PEKING UNIV
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