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Method for separating protoplast of tomato fruit

A protoplast and fruit technology, applied in the direction of plant cells, can solve the problems of fruit slicing, affecting fruit cell structure and function in situ non-destructive analysis, stabbing, etc., to achieve the effect of reducing dosage, improving yield and saving costs

Active Publication Date: 2017-05-24
INST OF BOTANY CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Therefore, microscopic observation of fruit cells was rare in the past, and the fruit was often sliced ​​or stabbed in the experiment, which greatly affected the in situ non-destructive analysis of fruit cell structure and function.

Method used

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  • Method for separating protoplast of tomato fruit
  • Method for separating protoplast of tomato fruit
  • Method for separating protoplast of tomato fruit

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0050] Embodiment 1, tomato fruit protoplast separation in red ripe stage

[0051] 1. Reagents required for separation

[0052] The enzyme activity of cellulase is 10u / mg (the enzyme activity under the β-1,4-D-glucan glucoside substrate is 10u / mg), which is a product of Yakult Japan;

[0053] The enzyme activity of pectinase is 5u / mg (the enzyme activity under pectin methoxy ester substrate is 5u / mg), is the product of Yakult Japan company;

[0054] The enzyme activity of the isolated enzyme is 3u / mg (the enzyme activity under the β-1,4-D-glucan glucoside substrate is 5u / mg), and it is a product of YakultJapan Company.

[0055] Enzyme liquid is made up of cellulase, pectinase, isolated enzyme and GKMMB solution, and the final concentration of cellulase is 1% (mass percentage), the final concentration of pectinase is 0.5% (mass percentage), the final concentration of isolated enzyme The concentration is 0.4% (mass percentage).

[0056] Above-mentioned GKMMB solution is by fi...

Embodiment 2

[0071] Embodiment 2, the tomato fruit protoplast separation of green maturity period

[0072] 1. Reagents required for separation

[0073] The enzyme activity of cellulase is 10u / mg (the enzyme activity under the β-1,4-D-glucan glucoside substrate is 10u / mg), which is a product of Yakult Japan;

[0074] The enzyme activity of pectinase is 5u / mg (the enzyme activity under pectin methoxy ester substrate is 5u / mg), is the product of Yakult Japan company;

[0075] The enzyme activity of the isolated enzyme is 3u / mg (the enzyme activity under the β-1,4-D-glucan glucoside substrate is 5u / mg), and it is a product of YakultJapan Company.

[0076] The enzyme liquid is the solution obtained by adding cellulase, pectinase and isolated enzyme to the GKMMB solution, the final concentration of cellulase is 1% (mass percentage), and the final concentration of pectinase is 0.4% (mass percentage) , The final concentration of the isolated enzyme is 0.5% (mass percentage).

[0077] Above-ment...

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Abstract

The invention discloses a method for separating protoplast of a tomato fruit. The method disclosed by the invention comprises the following steps: 1) performing enzymolysis on the tomato fruit by using an enzyme liquid, and collecting fruit tissue after enzymolysis, wherein the enzyme liquid comprises cellulose, pectinase and macerozyme; 2) performing incubation on the fruit tissue after enzymolysis in a PME (Pectin Methylesterase) solution, and separating the tomato fruit from protoplast, thereby obtaining tomato fruit protoplast. When the tomato fruit protoplast is separated by using the method disclosed by the invention, the yield of the protoplast can be remarkably increased, the use amounts of cellulose and the like can be reduced, the purpose of reducing the cost can be achieved, and an important tool for deep study on related gene expression modes of fruit development is provided, therefore, by using the method that enzymolysis treatment is combined with a gentle digestion technique for separating the tomato fruit protoplast, a basis is made for deep study on subcellular localization of related fruit development protein and gene expression modes, and high practical application values can be made.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for isolating tomato fruit protoplasts. Background technique [0002] As a fresh agricultural product, fruit provides rich nutrition for human health and is an indispensable component in people's dietary structure. Tomato, as a model organism for studying fruit ripening and disease resistance mechanism, is the largest vegetable crop widely planted in the world, and has important economic and scientific value. In recent years, the interpretation of the tomato genome sequence and the development of population genetics have enabled people to deepen our understanding of tomato from a genomic perspective. With the continuous deepening of functional genomics and molecular biology research, the spatiotemporal specificity and regulatory mode of functional genes related to fruit development process in cells have been widely concerned, and the analysis of subcellular localization of ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N5/04
Inventor 田世平陈彤秦国政徐勇
Owner INST OF BOTANY CHINESE ACAD OF SCI