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Method for Obtaining Differential Protein Map of Capsicum Anthers by Two-dimensional Electrophoresis

A technology of protein map and two-dimensional electrophoresis, which is applied in the field of acquisition of two-dimensional electrophoresis differential protein map of pepper anthers, can solve problems such as blanks, achieve the effect of large number, increased cleaning times, and uniform distribution

Active Publication Date: 2015-12-09
HANGZHOU ACAD OF AGRI SCI
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, the method of using two-dimensional electrophoresis to obtain protein maps has been applied to a certain extent in the field of biomedicine and some plant research (such as apple leaves), but it is almost blank in the research of peppers, especially anthers.

Method used

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  • Method for Obtaining Differential Protein Map of Capsicum Anthers by Two-dimensional Electrophoresis
  • Method for Obtaining Differential Protein Map of Capsicum Anthers by Two-dimensional Electrophoresis
  • Method for Obtaining Differential Protein Map of Capsicum Anthers by Two-dimensional Electrophoresis

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Experimental program
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Effect test

Embodiment 1

[0048] Embodiment 1, a kind of acquisition method of pepper anther two-dimensional electrophoresis difference protein atlas, the pepper material used is ' Hangjiao No. 1 ' of the cleavage type, get its anther; Carry out the following steps successively:

[0049] 1), protein extraction:

[0050] Include the following steps:

[0051] ①. Take 0.4g pepper anther and add 3ml liquid nitrogen, grind it and suspend it in 10ml solution containing 8% (v / v) trichloroacetic acid (prepared in acetone), and precipitate at -20°C for 10 hours;

[0052] ②. Centrifuge the product obtained in step ① (4°C, 13000r / min, 20min), and discard the supernatant;

[0053] ③. Suspend the precipitate obtained in step ② in 10ml of B-mercaptoethanol cold acetone solution; centrifuge (4°C, 13000r / min, 20min), repeat the suspension and centrifugation of the obtained precipitate 3 times;

[0054] The preparation method of the B-mercaptoethanol cold acetone solution is: mixing cold acetone and B-mercaptoethanol...

Embodiment 2

[0083] Embodiment 2, capsicum variety ' Bulgaria sharp pepper' anther substitutes the capsicum anther in embodiment 1, and all the other are the same as embodiment 1.

[0084] The final spectrum obtained is as figure 2 shown. according to figure 2 , we can draw the following conclusions respectively: for thicker fruit-shaped pepper anthers, the method of the present invention can also stably obtain high-quality protein expression maps. The protein map is clear, the number of protein spots is large, and the background is very clear.

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Abstract

The invention discloses a method for obtaining a two-dimensional electrophoresis difference protein map of chili pepper anther. The method comprises the following steps: (1) extracting protein by using the chili pepper anther; (2) carrying out first IPG isoelectric focusing electrophoresis; (3) balancing an adhesive tape; (4) transferring the adhesive tape and SDS-polyacrylamide gel electrophoresis; and (5) carrying out gel scanning and image analysis, so as to obtain the two-dimensional electrophoresis difference protein map of the chili pepper anther. By adopting the method disclosed by the invention, the two-dimensional electrophoresis map of chili pepper anther protein, which is clear in protein points, large in quantity, even in distribution and clear in background can be obtained, and the experiment result is stable. An important work basis is established for further research of chili pepper proteomics and molecular biology.

Description

technical field [0001] The invention relates to a method for obtaining a two-dimensional electrophoresis differential protein map of pepper anthers. Background technique [0002] Pepper (Chili Pepper) is native to the tropical regions of Central Latin America, and the country of origin is Mexico. It is an annual or perennial plant, its Vc content ranks first among vegetables, and it has high nutritional value. Capsicum is a frequently cross-pollinated plant with significant hybrid vigor. However, in hybrid breeding, it is time-consuming and labor-intensive due to emasculation of the female parent, and the purity of hybrid seeds is difficult to guarantee. The use of CMS three-line matching method to produce hybrid seeds can not only reduce the trouble of manual castration, reduce the cost of seed production, but also ensure the purity of hybrid seeds. At present, many capsicum cytoplasmic male sterile lines have been bred at home and abroad, and gradually applied to the pr...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N27/447G01N1/28
Inventor 吕晓菡柴伟国方献平
Owner HANGZHOU ACAD OF AGRI SCI
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