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Bidirectional electrophoresis method for total protein of jute root system

A two-dimensional electrophoresis, total protein technology, applied in the biological field, can solve problems such as failure to successfully prepare high-purity protein

Inactive Publication Date: 2012-10-10
NANJING AGRICULTURAL UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

[0004] However, because the plants contain a variety of secondary metabolites (pigments, phenols, quinones, etc.) produced under salinization conditions, many interferences make the core technology of proteome research - two-dimensional electrophoresis a challenge, and conventional methods cannot be successful. Prepare high-purity proteins. During electrophoresis, reasonable parameter settings can effectively desalt, focus, etc.

Method used

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  • Bidirectional electrophoresis method for total protein of jute root system
  • Bidirectional electrophoresis method for total protein of jute root system

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0051] US Beckmen high-speed low-temperature centrifuge, two-dimensional electrophoresis PROTEAN IIXi / XL Cell (US BIO-RAD)

[0052] 1) Collect indoor hydroponic jute roots, wash them with ultra-pure water, blot the water on the surface of the roots with filter paper, immediately put them into a liquid nitrogen tank for rapid freezing, and then store them in a refrigerator below -70°C in the laboratory for later use;

[0053]2) Put 2 g of jute root samples into an ice-bathed mortar and add liquid nitrogen to grind quickly. During the grinding process, add 0.2 g of polyvinylpyrrolidone until it is ground into a fine powder, and transfer it to a 50 ml centrifuge tube pre-cooled at -20°C;

[0054] 3) Add 3 times the volume of -20°C pre-cooled trichloroacetic acid extract, mix well, place at -20°C for 1.5h, and prepare the trichloroacetic acid extract: mass volume ratio 10% trichloroacetic acid TCA, mass volume ratio 0.07% dithiothreitol DTT, 1mM phenylmethylsulfonyl fluoride PMSF,...

Embodiment 2

[0081] US Beckmen high-speed low-temperature centrifuge, two-dimensional electrophoresis PROTEAN IIXi / XL Cell (US BIO-RAD)

[0082] 1) Collect the jute root system planted on the coastal tidal flats of Dafeng City, Jiangsu Province, clean it with ultrapure water, blot the surface water of the root system with filter paper, put it into a liquid nitrogen tank for rapid freezing, transport it back, and then store it in the laboratory -70 Store in refrigerator below ℃ for later use;

[0083] 2) Put 2 g of jute root samples into an ice-bathed mortar and add liquid nitrogen to grind quickly. During the grinding process, add 0.2 g of polyvinylpyrrolidone until it is ground into a fine powder, and transfer it to a 50 ml centrifuge tube pre-cooled at -20°C;

[0084] 3) Add 3 times the volume of -20°C pre-cooled trichloroacetic acid extract, mix well, place at -20°C for 1.5h, and prepare the trichloroacetic acid extract: mass volume ratio 10% trichloroacetic acid TCA, mass volume ratio ...

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Abstract

The invention relates to a dimensional electrophoresis method for a total protein of a jute root system, belonging to the field of biotechnology. The total protein of the jute root system capable of existing in the coastal beach is separated by the dimensional electrophoresis technique, and by adopting the technical scheme, the jute root system existing in the coastal beach is tested with good effect. At present, repeated experiments prove that the jute root system is separated by the dimensional electrophoresis method to obtain more protein points; and after detected by PDQuest software, theprotein points are more than 1100, the map is clear without transverse and longitudinal grains, the protein points are circular, and the repeatability is good, thus the invention is a dimensional electrophoresis method suitable for the analysis of the total proteomics of the jute root system.

Description

technical field [0001] The invention relates to a two-dimensional electrophoresis method for total jute root protein, belonging to the field of biotechnology. It is the proteome separation of jute roots that can grow in a large area on coastal beaches, and this method can be directly applied to the research, development and utilization of highly salt-tolerant plants that can survive in coastal beaches. Background technique [0002] Stress resistance is a countermeasure that plants acquire in order to adapt to adverse environments in the process of systematic evolution. In the living environment of plants, some abiotic factors stress will have a serious impact on the growth and survival of plants. In order to cope with the adverse effects of the external environment, plants have formed a complete defense mechanism during the evolution process. In order to analyze its defense mechanism, it is necessary to isolate some proteins related to adversity. [0003] Jute (Orchorus ol...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N27/447G01N1/28G01N1/30
Inventor 麻浩马洪雨王显生何小玲
Owner NANJING AGRICULTURAL UNIVERSITY
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