Plant pollen tube cell calcium ion ultrastructure positioning method

A plant pollen and ultrastructure technology is applied in the preparation of test samples, material analysis using radiation, etc. It can solve the problems of difficult to find the central axis and longitudinal length of the pollen tube, avoid non-specific background pollution, The effect of prolonging the rinsing process and reducing contamination

Inactive Publication Date: 2015-07-22
CROP RES INST SHANDONG ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

However, pollen and its germinated pollen tubes are unicellular, and the diameter of the pollen tube is only 20-40 μm, and the length is long.

Method used

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  • Plant pollen tube cell calcium ion ultrastructure positioning method
  • Plant pollen tube cell calcium ion ultrastructure positioning method

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Experimental program
Comparison scheme
Effect test

Embodiment 1

[0037] (1) Preparation of the solution:

[0038] The preparation of 0.2mol / L potassium phosphate buffer solution (pH7.8): stoste I: with 2.72g KH 2 PO 4 Dissolve in 100ml ddH 2 O; stock solution Ⅱ: 4.56g K 2 HPO 4 ·3H 2 O dissolved in 100ml ddH 2 O, then the stock solutions I and II were mixed in a volume ratio of 13:87;

[0039] 2.5% glutaraldehyde fixative solution (pH7.8): Take 90ml of the newly prepared above-mentioned 0.2mol / L potassium phosphate buffer solution, add 2g potassium pyroantimonate, heat to dissolve; Dialdehyde solution 10ml, sucrose 1.5g, tannic acid 0.2g, Triton X-1000.5ml, with ddH 2 O is settled to 100ml, and the final concentration of potassium pyroantimonate is 2%;

[0040] Preparation of washing buffer: take 50ml 0.2mol / L potassium phosphate buffer (pH7.8) and add 50ml ddH 2 O, then add 2g of potassium pyroantimonate, heat to dissolve, after filtering, use ddH 2 O is fixed to 100ml;

[0041] 1% osmic acid (OsO 4 ) Preparation of fixative so...

Embodiment 2

[0054] (1) Preparation of the solution:

[0055] The preparation of 0.2mol / L potassium phosphate buffer solution (pH7.8): stoste I: with 2.72g KH 2 PO 4 Dissolve in 100ml ddH 2 O; stock solution Ⅱ: 4.56g K 2 HPO 4 ·3H 2 O dissolved in 100ml ddH 2 O, then the stock solutions I and II were mixed in a volume ratio of 13:87;

[0056] 2.5% glutaraldehyde fixative solution (pH7.8): Take 90ml of the newly prepared above-mentioned 0.2mol / L potassium phosphate buffer solution, add 2g potassium pyroantimonate, heat to dissolve; Dialdehyde solution 10ml, sucrose 1.5g, tannic acid 0.2g, Triton X-1000.5ml, with ddH 2 O is settled to 100ml, and the final concentration of potassium pyroantimonate is 2%;

[0057] Preparation of washing buffer: add 50ml ddH to 50ml 0.2mol / L potassium phosphate buffer (pH7.8) 2 O, then add 2g of potassium pyroantimonate, heat to dissolve, after filtering, use ddH 2 O is fixed to 100ml;

[0058] 1% osmic acid (OsO 4 ) Preparation of fixative solution...

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Abstract

The invention discloses a plant pollen tube cell calcium ion ultrastructure positioning method which mainly comprises the following steps: (1) preparing a quantitative immobilizing liquid and washing liquid; (2) immobilizing and washing a material; (3) immobilizing and washing secondarily; (4) dehydrating and transiting; (5) carrying out resin permeating, embedding and polymerizing; (6) directly observing by using an electron microscope without dyeing the material. Due to the adoption of the method, the problems that cell turgor pressure needs to be adjusted, abnormal reaction can be caused between potassium pyroantimonate and buffer liquid, calcium pyroantimonate precipitation displacement or dilution can be caused by material immobilization and aftertreatment, Ca<2+> precipitate observation can be affected because of inappropriate dyeing, and the like can be solved, and organelle can be clearly distinguished without dyeing completely.

Description

technical field [0001] The invention belongs to the technical field of ultrastructure observation and preparation of plant cells and chemical composition positioning, and particularly relates to a Ca in plant pollen tube cells. 2+ Qualitative and positional observation techniques. Background technique [0002] Inorganic ions play an important physiological role in plant cells, such as regulating enzyme activity, serving as a component of cell structure, regulating intracellular ion balance, cell osmotic regulation, and stomatal movement. Ca 2+ It is not only an essential element for plant growth and development, but also the most important second messenger that regulates cell physiological responses. Different developmental signals and external stimuli, including light, mechanical interference, abiotic stress, and pathogenic elicitors, can all cause Ca 2+ Level changes (Farnklin-tong, 1999). In the normal development process of plants and in response to external stimuli s...

Claims

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Application Information

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IPC IPC(8): G01N23/04G01N1/28
Inventor 孔令安林金星王钦丽王晓华谢燕
Owner CROP RES INST SHANDONG ACAD OF AGRI SCI
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