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Preparation method of pyrus betulaefolia protoplast

A technology for separating protoplasts and plasmodium, which is applied in the field of cell biology and biology to achieve the effect of overcoming the difficulty of separation

Inactive Publication Date: 2017-07-21
JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

At present, there is no report about the separation research of Du pear protoplasts, and a kind of simple and efficient separation and preparation method of Du pear protoplasts that the present invention attempts to establish will be the basis for Du pear cell biology research and Du pear single-cell genetic transformation system. create basic conditions

Method used

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  • Preparation method of pyrus betulaefolia protoplast

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0034] (1) choose the young leaves of the 60, 75, 90d growing and vigorous seedling ages of Du pear seedlings as processing material, put into ultra-clean workbench after washing with tap water, sterilize with 70% dehydrated alcohol, rinse with aseptic water and stand-by;

[0035] (2) Treat the material after step (1) in a dark environment at room temperature for 0.5h, and cut it into 0.5-1mm wide fine leaf shreds along the vein direction with a blade after treatment;

[0036] (3) Weigh 1 g of the finely chopped leaves and put them into 20 ml of plasmolysis treatment solution, then place them in a dark environment at room temperature, and treat them in the dark for 1 hour;

[0037](4) Take 1g of the above-mentioned thin leaf silks that have undergone plasmolysis treatment, put them into a pre-prepared petri dish containing 10ml of enzymatic hydrolysis solution, and then place it in an environment of 25-28°C, and enzymatically hydrolyze it for 8 hours in the dark. , when the en...

Embodiment 2

[0043] (1) choose the young leaves of the 60, 75, 90d growing and vigorous seedling ages of Du pear seedlings as processing material, put into ultra-clean workbench after washing with tap water, sterilize with 70% dehydrated alcohol, rinse with aseptic water and stand-by;

[0044] (2) Treat the material treated in step (1) for 1 hour at room temperature in a dark environment, and cut it into 0.5-1 mm wide fine leaf silks along the vein direction with a blade after treatment;

[0045] (3) Weigh 1 g of the finely cut shredded leaves and put them into 20 ml of plasmolysis treatment solution, then place them in a dark environment at room temperature, and treat them in the dark for 3 hours;

[0046] (4) Take 1 g of the above-mentioned thin leaf silks that have undergone plasmolysis treatment, put them into a pre-prepared petri dish containing 10 ml of enzymatic hydrolysis solution, and then place them in an environment of 25-28 ° C, corresponding to the light-proof enzyme Decompose...

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Abstract

The invention belongs to the technical fields of cytobiology and biology, and discloses a preparation method of pyrus betulaefolia protoplast. According to the method, through the pretreatment of a pyrus betulaefolia material and separation treatment of cytoplasm and cytoderm, the mesophyll tissue of pyrus betulaefolia is subjected to enzymolysis through the combination of cellulase, macerase and pectinase, the preparation of pyrus betulaefolia protoplast is effectively realized, the problem that the pyrus betulaefolia protoplast is difficult to separate is overcome, and the preparation method of the pyrus betulaefolia protoplast is effectively created. The method breaks through the bottleneck of genetic transformation of pyrus betulaefolia, establishes a pyrus betulaefolia cell engineering technical system, and provides a technical base for the genetic transformation of the pyrus betulaefolia protoplast, the transient expression examination of gene and the fusion of the protoplast.

Description

technical field [0001] The invention belongs to the field of cell biology and biotechnology, in particular to a method for preparing pear protoplasts. Background technique [0002] Due to the absence of cell walls, protoplasts have their own unique characteristics relative to tissues, organs or individuals: first, they can eliminate the interaction between cells and examine the totipotency of a cell alone; second, protoplasts are a homogeneous A large number of protoplasts may be obtained in a short period of time for a group with a high degree of sex. Studies have shown that protoplasts can not only be used as a single-cell system to study cell wall regeneration, cell division and differentiation, uptake of organelles, virus infection mechanism, membrane permeability and ion transport, but also an ideal material for basic theoretical research; it is also an ideal material for protoplast culture. Basis for fusion with protoplasts. [0003] In recent years, plant protoplast...

Claims

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Application Information

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IPC IPC(8): C12N5/04
CPCC12N5/04
Inventor 唐君常有宏蔺经李晓刚
Owner JIANGSU ACADEMY OF AGRICULTURAL SCIENCES
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