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Method for staining iron element in plants

A dyeing method and iron element technology, which is applied in the field of plant biology, can solve the problems of time-consuming and laborious, and achieve the effect of simple operation and short cycle

Inactive Publication Date: 2013-07-17
SOUTH CHINA BOTANICAL GARDEN CHINESE ACADEMY OF SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Whether chloral hydrate clearing agent can be used in Perls iron staining to replace time-consuming and laborious sectioning has not been reported yet

Method used

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  • Method for staining iron element in plants
  • Method for staining iron element in plants
  • Method for staining iron element in plants

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] Example 1: Iron staining and microscope observation in Arabidopsis seeds

[0030] (1) In order to facilitate the penetration of Perls staining solution, cut off part of the Arabidopsis seed coat with a sharp blade.

[0031] (2) Soak the seeds treated in step (1) in freshly prepared Perls staining solution, and stain for 30 minutes;

[0032] Preparation of Perls staining solution: 4% potassium ferrocyanide and 4% hydrochloric acid are mixed in equal amounts, matured for 15 minutes and set aside;

[0033] (3) Rinse with deionized water for 3 times;

[0034] (4) Put the rinsed seeds in the chloral hydrate test solution, and make them transparent for more than 8 hours, or overnight, until the seeds are completely transparent;

[0035] Preparation of chloral hydrate test solution: 50g of chloral hydrate, 15ml of water and 10ml of glycerin, dissolved to obtain;

[0036] (5) Take an appropriate amount of chloral hydrate test solution transparent agent and drop the seeds tre...

Embodiment 2

[0038] Embodiment 2: iron staining and microscope observation in rice embryo

[0039] (1) Dissect the embryo from the rice seed with a sharp blade and a dissecting needle

[0040] (2) Immerse the dissected embryos in freshly prepared Perls staining solution and stain for 30 minutes;

[0041] Preparation of Perls staining solution: 4% potassium ferrocyanide and 4% hydrochloric acid are mixed in equal amounts, matured for 15 minutes and set aside;

[0042] (3) Rinse with deionized water for 3 times;

[0043] (4) Place the rinsed embryos in the chloral hydrate test solution, and make them transparent for more than 8 hours, or overnight, until the embryos are completely transparent;

[0044] Preparation of chloral hydrate test solution: 50g of chloral hydrate, 15ml of water and 10ml of glycerin, dissolved to obtain;

[0045] (5) Place the embryos treated in step (4) on a glass slide, cut the embryos into 2 with a sharp blade, cover with a cover glass, and press slightly. The f...

Embodiment 3

[0046] Embodiment 3: comparative test

[0047] A number of mature seeds of Arabidopsis thaliana were selected and randomly divided into two parts. One part was dyed by the method for iron element dyeing in plants of the present invention. See Example 1 for specific steps.

[0048] One section was stained with Perls-DAB, the specific steps are as follows: (1) Put the Arabidopsis seeds in 10% neutral formalin (0.01M PBS, PH=7.4) to fix for more than 10h, rinse with 0.01MPBS 3 times, 30 minutes each time; (2) Serial alcohol dehydration, 1-2 hours per level; (3) TO transparent agent transparent 3 times, 2 hours each time; (4) Permeable wax, broken wax and TO transparent agent 1:1 , Overnight at 40 degrees, 60 degrees, change pure wax twice, 4 hours each time; (5) Embed in paraffin, slice with American AO microtome, thickness 6-8um, bake slices at 40 degrees overnight; (6) Dewax and rehydrate : TO clearing agent 2 times, TO clearing agent and absolute ethanol 1:1, 5 minutes each t...

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Abstract

The invention discloses a method for staining the iron element in plants. The method comprises steps of: cutting a plant material into small pieces; soaking the small pieces into a Perls staining fluid to be stained; rinsing the stained small plant material pieces with deionized water; putting the rinsed small plant material pieces into a chloral hydrate test solution until the small plant material pieces are completely transparent; and taking out the small plant material pieces so as to obtain the materials with stained iron element. According to the method, a Prussian blue Perls iron staining method and a chloral hydrate transparency method are organically combined together, the internal iron distribution of the plant can be integrally displayed under the condition of keeping the plant material complete to the greatest extent; the method has a short period, and a result is generally obtained after a week through adopting a slice method, but can be obtained in a day through utilizing the method; and the method has the advantages of simplicity in operation and no adoption of expensive equipment.

Description

Technical field: [0001] The invention belongs to the field of plant biotechnology, and in particular relates to a method for dyeing iron elements in plants. Background technique: [0002] Prussian blue (Perls) staining is an ancient and sensitive staining method for iron display. The principle of the reaction is that high-valent iron in the tissue reacts with potassium ferrocyanide to form an insoluble blue precipitate (Prussian blue). Because of its simple operation, low cost, and good specificity, it is now the most commonly used method in laboratories to display iron distribution in organisms (Meguro, Asano et al.2007). At present, in the study of plant iron metabolism, the method of displaying iron distribution is to add Perls staining on the basis of sections (Green and Rogers2004, Roschzttardtz, Conejero et al.2009, Zhang, Xu et al.2012). This method not only cannot show the distribution of iron in the plant as a whole, but also has a long test period (about one week...

Claims

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Application Information

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IPC IPC(8): G01N1/30
Inventor 叶宏刘培卫秦璐王美欢梁雪娇
Owner SOUTH CHINA BOTANICAL GARDEN CHINESE ACADEMY OF SCI
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