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Immobilized modification method for plant suspension cells

A technique of suspending cells and activators, applied in biochemical equipment and methods, measurement/inspection of microorganisms, immobilization on or in inorganic carriers, etc., which can solve the problem of poor adherence effect, large force, and difficulty in real-time reflection The true state of cell structure and function, etc.

Inactive Publication Date: 2012-05-02
INST OF CHEM CHINESE ACAD OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0003] At present, many literatures have reported the fixation methods of suspension cells, such as poly-lysine cross-linking, micropipette mouth capture, small-pore filter membrane interception method, etc., but these methods have defects to varying degrees.
In animal cell experiments, polylysine is only used as an auxiliary cross-linking method to enhance the adhesion effect of cells that are not easily adhered to or semi-adhered to the wall. It is used to fix suspended cells, and the efficiency is low and the adhesion effect not good
However, methods such as micropipette capture and small-pore membrane interception exert too much force on the cell surface, especially for plant cell protoplasts, destroying the normal physiological state of biological cells, making it difficult to reflect cell structure and function in real time the real situation of

Method used

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  • Immobilized modification method for plant suspension cells
  • Immobilized modification method for plant suspension cells
  • Immobilized modification method for plant suspension cells

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0061] Example 1. Obtainment and application of plant suspension cells fixed and cross-linked to the substrate

[0062] 1. Obtaining plant suspension cells immobilized and cross-linked to the substrate

[0063] 1. Activate plant cells and activate substrate materials

[0064] 1) Activate plant suspension cells

[0065] (1) Take 1 ml of tobacco suspension cells (Nicotiana tobaccocum L. cv. Bright Yellow, BY-2) in good growth state, and add it to an Erlenmeyer flask containing 20 ml of medium. Protected from light and placed in a shaker at 26°C with a rotation speed of 120rpm, cultured for 3 days to obtain a BY-2 cell solution (which can be used as a growing BY-2 cell for later experiments, wherein the concentration of BY-2 cells is 2.36×10 3 pieces / ml.

[0066] (2) Take 4ml of BY-2 cells in step (1), transfer to a 15ml centrifuge tube, let it settle to remove the supernatant, then add 2ml of culture medium, gently pipette evenly, and transfer to a 60mm culture dish; Then we...

Embodiment 2

[0093] Example 2, the acquisition of plant suspension cell protoplasts fixed to the substrate by cross-linking

[0094] According to the same method and steps as 1-2 of step 1 of Example 1, the plant suspension cells were immobilized and modified, and only the BY-2 cells used were replaced with the protoplasts of BY-2 cells to obtain the fixed on the glass. BY-2 protoplasts (glass slides immobilized with BY-2 protoplasts).

[0095] The BY-2 protoplasts fixed on the glass (glass slides with BY-2 protoplasts fixed) were detected in step 1 of Example 1, and the normal growing protoplasts were used as contrast;

[0096] The result is as Figure 4 As shown, A, B, and C are BY-2 protoplasts growing in suspension, and D, E, and F are images of BY-2 protoplasts fixed on glass. Among them, A and D are the bright field images of the negative control, B and E are the fluorescence images stained by FDA, and C and F are the superimposed images of the bright field images and the fluoresce...

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Abstract

The invention discloses an immobilized modification method for plant suspension cells. The method for immobilizing the suspension cells specifically comprises the following steps of: crosslinking the suspension cells with substrate materials: 1) activating the suspension cells and the substrate materials respectively to obtain activated suspension cells and activated substrate materials; and 2) subjecting the activated suspension cells and the activated substrate materials obtained in the step 1) to a reaction and thus obtaining the suspension cells immobilized on the substrate materials. Theexperiments of the immobilized modification method show that the invention belongs to an immobilized modification method for suspension growth cells. Compared with the similar technology, the immobilized modification method has the advantages of simple operation, high cell adherence, normal biological activity, near physiological environment and the like. By the immobilized modification method, various suspension growth biological cells can be immobilized.

Description

technical field [0001] The invention relates to the field of biotechnology, in particular to a method for immobilizing and modifying plant suspension cells. Background technique [0002] Atomic force microscopy (AFM) has been widely used in many fields because of its high spatial resolution and mechanical sensitivity, and it has also been greatly developed in the field of biology. In recent years, the high-resolution imaging and single-molecule force measurement of AFM have made in-depth research on the ultrastructure and dynamic structure of biomacromolecules and cell membranes, as well as the interaction forces between DNA, antibody-antigen, drugs and biomolecules. Research. However, most of the research on AFM on living cells is limited to animal adherent cells, plant suspension cells and their protoplasts due to the special state of suspension growth, and the lack of a suitable method for fixing suspension cells, making the living cell level The research methods on the...

Claims

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Application Information

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IPC IPC(8): C12N11/14C12Q1/02
Inventor 方晓红张雪洁刘巧玲师晓丽
Owner INST OF CHEM CHINESE ACAD OF SCI
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