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A kind of dyeing solution and method for rapid dyeing of pollen of cruciferous crops

A rapid dyeing technology for cruciferous crops, applied in the preparation of test samples, scientific instruments, sampling, etc., can solve the problems of tedious, time-consuming, labor-consuming, etc., achieve good repeatability, obvious contrast of nuclear staining, and reduce Effects of Environmental Pollution

Active Publication Date: 2021-03-23
GUANGDONG KING ZUO AGRI SCI & TECH CO LTD +1
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Aiming at the deficiencies of the prior art, the present invention aims to provide a dyeing solution and method for rapid dyeing of pollen of cruciferous crops, which solves the problems of cumbersome, time-consuming and labor-consuming existing pollen fluorescent dyeing methods, and at the same time improves the fluorescence Stability and reproducibility of staining

Method used

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  • A kind of dyeing solution and method for rapid dyeing of pollen of cruciferous crops
  • A kind of dyeing solution and method for rapid dyeing of pollen of cruciferous crops
  • A kind of dyeing solution and method for rapid dyeing of pollen of cruciferous crops

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Effect test

Embodiment 1

[0025] Embodiment 1: Fluorescent staining method of mature pollen of open flowers of rapeseed

[0026] 1. Preparation of staining solution: first dissolve Hoechst33258 fluorescent dye and sucrose in phosphate buffered saline (PBS, pH=7.4) to prepare solution 1 of Hoechst33258 concentration 20 μg / L and sucrose concentration 10wt%, and then prepare ethanol 60% ( v / v), ethyl acetate acetic acid 10% (v / v), glycerol 30% (v / v) solution two, solution one and solution two were mixed and shaken by volume ratio 1:1 to obtain the staining solution, and placed side by side Store in a refrigerator at 4°C, protected from light.

[0027] 2. Staining: Use a micropipette to absorb 15 μL of the staining solution obtained in step 1 and drop it on the glass slide, then pick the open flowers that need to be stained and observed from the inflorescence, and use tweezers to pick one anther and put it directly on the glass slide. In the staining solution, gently pinch the anthers to release an approp...

Embodiment 2

[0030] Embodiment 2: the fluorescent staining method of Chinese cabbage flower bud pollen

[0031] 1. Preparation of staining solution: first dissolve Hoechst33258 fluorescent dye and sucrose in phosphate buffered saline (PBS, pH=7.4) to prepare solution 1 of Hoechst33258 concentration 30 μg / L and sucrose concentration 8wt%, and then prepare ethanol containing 70% ( v / v), ethyl acetate acetic acid 10% (v / v), glycerol 20% (v / v) solution two, solution one and solution two were mixed and shaken by volume ratio 1:1 to obtain the staining solution, placed in Store in a refrigerator at 4°C, protected from light.

[0032] 2. Staining: Use a micropipette to absorb 17 μL of the staining solution prepared in step 1 and drop it on the glass slide, then pick the flower buds that need to be stained and observed from the inflorescence, and use tweezers to peel off one anther and put it directly on the glass slide Gently pinch the anthers in the staining solution to scatter an appropriate a...

Embodiment 3

[0035] Embodiment 3: the fluorescent staining method of cabbage flower bud pollen

[0036] 1. Preparation of staining solution: first dissolve Hoechst33258 fluorescent dye and sucrose in phosphate buffered saline (PBS, pH=7.4) to prepare solution 1 with Hoechst33258 concentration 40 μg / L and sucrose concentration 9 wt%, and then prepare ethanol containing 65% ( v / v), ethyl acetate acetic acid 10% (v / v), glycerol 25% (v / v) solution two, solution one and solution two were mixed and shaken by volume ratio 1:1 to obtain the staining solution, juxtaposed Store in a refrigerator at 4°C, protected from light.

[0037] 2. Staining: Use a micropipette to absorb 20 μL of the staining solution obtained in step 1 and drop it on the glass slide, then pick the flower buds that need to be stained and observed from the inflorescence, and use tweezers to pick 2 anthers and put them directly on the glass slide. In the staining solution, gently pinch the anthers to release an appropriate amount...

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Abstract

The invention discloses a fast fluorescent staining method of cruciferae crop pollen. The method comprises the following steps of S1, staining liquid preparation: firstly, dissolving Hoechst33258 fluorescent dye and cane sugar into a PBS (phosphate buffer solution); preparing a first solution with the Hoechst33258 concentration being 20 to 40mug / L and the cane sugar concentration being 8 to 10 weight percent; then, preparing a second solution containing 60 to 70 percent (v / v) of ethanol, 10 percent (v / v) of ethyl acetate and 30 to 20 percent (v / v) of glycerol; mixing and uniformly mixing the first solution and the second solution according to a volume ratio of 1:1 to obtain staining liquid; performing low-temperature light avoiding storage; S2, staining: taking staining liquid prepared inthe steps S1 on a glass slide; then, picking blossoming flowers or flower buds to be stained and observed from the inflorescence; picking the anther by tweezers to be directly put into staining liquidon the glass slide; slightly pinching the anther; spreading out pollen; taking out sundries; performing smearing so that the pollen and the staining liquid are uniformly mixed; then, cover glass is covered. The problems of complexity, time waste and labor waste of the existing pollen fluorescent staining method can be solved; meanwhile, the stability and the repeatability of the fluorescent staining method are improved.

Description

technical field [0001] The invention relates to a pollen fluorescent dyeing method, in particular to a dyeing solution and method for rapid dyeing of cruciferous crop pollen. Background technique [0002] Plant pollen wall is a special cell wall mainly composed of sporopollenin and cellulose, which is composed of two layers: outer wall and inner wall. Among them, the outer wall is divided into inner and outer layers. The outer layer of the outer wall is honeycomb or mesh-like, while the inner layer of the outer wall is flat. The main component of the outer wall is sporopollenin. The inner wall structure is relatively simple, mainly containing cellulose, pectin and protein. Because the structure of the plant pollen wall is more complex than that of the general plant cell wall, and it is compact and thick, especially the mature pollen grains, which have extremely poor permeability and light transmission. Therefore, most plant pollen is difficult to observe under a microscope ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N1/30
CPCG01N1/30G01N2001/302
Inventor 李智军曾晶谭铭许谢景
Owner GUANGDONG KING ZUO AGRI SCI & TECH CO LTD
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