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Fluorescent staining method for observing plant microstructure

A technology of microstructure and fluorescent staining, which is applied in the field of plant tissue observation, can solve the problems of shortening the preparation time and achieve good economic and social benefits, simple method and low cost

Inactive Publication Date: 2016-07-06
HENAN UNIV OF CHINESE MEDICINE
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It not only greatly shortens the time required for sample preparation, but also can obtain three-dimensional images, and has been widely used in botanical research. However, there is no effective method on how to apply fluorescent dyeing to the observation of plant microstructure. public report

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0027] In concrete implementation, the present invention is realized by the following steps:

[0028] 1. Take materials

[0029] Take pollen grains, embryo sacs, styles or ovules of plants, or take leaves, roots or stems and cut them into small pieces or segments of 2-3mm in length and width;

[0030] 2. fixed

[0031] At room temperature, place the plant material in a glass container filled with a fixative solution made of paraformaldehyde or methanol with a mass concentration of 4% prepared by PBS buffer solution with a pH of 8.0, and fix it for 25-35min. When fixing the leaves , use methanol to remove chlorophyll and dissolve the cuticle;

[0032] 3. First rinse

[0033] Take out the plant material from the fixative solution and rinse with PBS buffer solution of pH 8.0 for 12-18min;

[0034] 4. Cell Wall Staining

[0035] Put the plant material after the first rinsing in the circular concave of the double-concave glass slide, add 0.03-0.05ml of DMSO with a mass concent...

Embodiment 2

[0045] In concrete implementation, the present invention is realized by the following steps:

[0046] 1. Take materials

[0047] Take pollen grains, embryo sacs, styles or ovules of plants, or take leaves, roots or stems and cut them into small pieces or segments of 1-5mm in length and width;

[0048] 2. Fixed

[0049] At room temperature, place the plant material in a glass container filled with a fixative solution made of paraformaldehyde or methanol with a mass concentration of 4% prepared by PBS buffer solution with a pH of 8.0, and fix it for 30 minutes. When fixing the leaves, it is necessary to Use methanol to remove chlorophyll and dissolve the cuticle;

[0050] 3. First rinse

[0051] Take out the plant material from the fixative, rinse with PBS buffer solution of pH 8.0 for 15min;

[0052] 4. Cell Wall Staining

[0053] The plant material after the first rinsing was placed in the concavity of the double-concave glass slide, and 0.06ml of DMSO with a mass concent...

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Abstract

The invention relates to a fluorescent dyeing method for observing the microstructure of plants, which can effectively solve the problem of rapid film production and observation of the microstructure of plants. , the plant material was fixed in paraformaldehyde or methanol with a mass concentration of 4% configured by pH 8.0 PBS buffer solution for 20-40min, rinsed with pH 8.0 PBS buffer solution for 10-20min, and prepared by DMSO, Triton X-100 and fluorescent whitening agent 220 make staining solution, stain at room temperature for 10-60min, then rinse with PBS buffer for 10-15min, add DAPI solution dropwise for fluorescent staining of nucleic acid in cells, rinse with distilled water, and then add dropwise Glycerin, add a cover glass, observe and take pictures with a laser confocal microscope, the method of the present invention is simple, does not need a microtome, and the production time is short, and the three-dimensional structure of the plant can be observed, which is suitable for the observation of cells or tissues of various plants, clear , accurate, low cost and good effect.

Description

technical field [0001] The invention relates to plant tissue observation, in particular to a fluorescent dyeing method for plant microstructure observation. Background technique [0002] Microscopic technology is an important means of studying plant science. At present, the research on the microstructure of plant cells and tissues is still dominated by optical microscopy, mainly using paraffin section and ultrathin section technology, but the preparation method is complicated and takes a long time. , cannot be quickly produced, and has high requirements for the operation of the experiment, requiring corresponding slicers and other equipment, and finally obtains two-dimensional images, which limits its wide use. [0003] Laser confocal microscopy can observe thicker, translucent or even opaque plant materials, and can detect the distribution of fluorescent substances in cells and tissues more sensitively. Information, after being processed by computer, its three-dimensional ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): G01N1/30
CPCG01N1/30
Inventor 刘孟奇王小巧赵乐罗晓代丽萍陈随清
Owner HENAN UNIV OF CHINESE MEDICINE
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