Method for Detecting Cell Number of Arabidopsis Leaf
An Arabidopsis thaliana and leaf technology is applied in the field of accurately and objectively detecting the number of Arabidopsis thaliana leaf cells, which can solve the problems of large statistical errors, incomplete cell separation, uneven distribution of protoplasts, etc., and achieves the effect of convenient operation.
Active Publication Date: 2016-05-04
HUNAN AGRICULTURAL UNIV
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Problems solved by technology
[0002] At present, counting the number of cells in plant leaves is mainly through the observation of the cell counting plate. Under the same sample processing conditions, the enzymatic hydrolysis time is difficult to control. If the enzymatic hydrolysis time is too short, the cells will be incompletely separated and there will be more adhesions. If the enzymatic hydrolysis time is too long , protoplasts are easy to rupture, and the proportion of complete cells obtained is not high, resulting in large statistical errors
When sampling and counting, the protoplasts are unevenly distributed in the enzyme solution, which makes the counting repeatability low and subjectivity strong
Method used
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Embodiment
[0025] Embodiment detects the method for Arabidopsis leaf cell number
[0026] 1 Test materials and reagents
[0027] 1.1 Test material
[0028] Arabidopsis Columbia ecotype wild-type plants (WT) and rfc3-1 mutant were used as test materials.
[0029] 1.2 Main Drugs and Reagents
[0030] Propidium iodide (PI), spermine tetrahydrochloride were purchased from Sigma Company; MES (MethylMethanesulfonate, methyl methanesulfonate), cellulase, isolated enzyme and β-mercaptoethanol were purchased from Pengcheng Biological Co., Ltd., other conventional chemical Reagents were purchased from Sinopharm Chemical Reagent Co., Ltd.
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Abstract
The invention provides a method for detecting the cell number of an arabidopsis leaf. The method comprises the steps of enzymolysis of the plant leaf; cell lysis and cell nucleus collection; cell nucleus dyeing and counting, and the like. The cell number of the arabidopsis leaf is determined by detecting the cell nucleus number with a flow cytometry. Compared with a manner that the cell number of the plant leaf is observed and counted by a cell counting plate, the method is capable of counting the cell number of the whole plant leaf, and has the advantages of being convenient to operate, objective and precise.
Description
technical field [0001] The invention relates to the field of biotechnology, in particular to a method for accurately and objectively detecting the number of Arabidopsis leaf cells. Background technique [0002] At present, counting the number of cells in plant leaves is mainly through the observation of the cell counting plate. Under the same sample processing conditions, the enzymatic hydrolysis time is difficult to control. If the enzymatic hydrolysis time is too short, the cells will be incompletely separated and there will be more adhesions. If the enzymatic hydrolysis time is too long , protoplasts are easy to rupture, and the proportion of complete cells obtained is not high, resulting in large statistical errors. When sampling and counting, the distribution of protoplasts in the enzyme solution is not uniform, so that the repeatability of counting is not high and the subjectivity is strong. Contents of the invention [0003] The purpose of the present invention is ...
Claims
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IPC IPC(8): G01N15/10
Inventor 夏石头崔看彭克勤肖浪涛
Owner HUNAN AGRICULTURAL UNIV