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Method for extracting total plant proteins

A technology of plant total protein and protein, applied in the preparation method of peptides, chemical instruments and methods, preparation of test samples, etc., can solve the problems of non-modification, difficulty in obtaining non-pollution, etc.

Inactive Publication Date: 2013-02-13
BEIJING UNIV OF AGRI
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Problems solved by technology

[0003] Especially for plant tissues, which not only contain a large number of cell wall structures, proteases and oxidases, but also contain a large number of polysaccharides and secondary metabolites (such as lipids, phenols, starch, etc.), these unfavorable factors make it difficult to obtain Non-polluting, non-modified, complete and high-quality plant cell protein

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  • Method for extracting total plant proteins
  • Method for extracting total plant proteins
  • Method for extracting total plant proteins

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Embodiment 1

[0027] Example 1, Extraction of Tomato Total Protein (the improved phenol-methanol method protected by the present invention)

[0028] Preparation method of protein extraction buffer: Dissolve 500mmol Tris-HCl, 50mmol EDTA, 700mmol sucrose and 100mmol KCl in water and dilute to 1L with water, adjust pH to 8.0, store at 4°C, add β-mercaptoethanol and each Protease, so that the concentration of β-mercaptoethanol is 2% (volume percentage), and the concentration of chymotrypsin is 1.5μg·ml -1 , The concentration of thermolysin is 0.8g·ml -1 , the concentration of papain is 1μg·ml -1 , the concentration of pronase is 1.5 μg·ml -1 , the concentration of trypsin is 15μg·ml -1 , The concentration of trypsin is 0.2μg·ml -1 .

[0029] Preparation method of protein precipitation solution: dissolve ammonium acetate in methanol, and dilute to 0.1mol L with methanol -1 , stored at -20°C.

[0030] Preparation method of protein lysate: dissolve 6mol urea, 2mol thiourea, 20mmol CHAPS an...

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Abstract

The invention discloses a method for extracting total plant proteins, in particular to a method for extracting the total proteins in the phloem of tomato plants. The method includes the following steps: (1) adding crosslinked polyvinylpyrrolidone (PVPP) and quartz sand in plant materials, and grinding the mixture into powder through liquid nitrogen; (2) adding protein extraction buffer solution into the powder, oscillating to homogenate the slurry, and then adding Tris saturated phenol for extracting to obtain an organic phase; (3) evenly mixing the organic phase with the protein sedimentation liquid, standing, and centrifuging to collect the sediments; and (4) sequentially performing methanol cleaning, acetone cleaning, freezing and drying on the sediments to obtain the total plant proteins. The main improvement of the method for extracting total plant proteins is that: (1) the PVPP is added during the grinding and smashing of samples to absorb phenolic substances, and the PVPP can be removed through centrifugal after the phenolic substances are adsorbed, so that the downstream analysis process cannot be affected; and (2) a plurality of protein enzymes are added during the extracting, so that the activity and natural characteristics of the proteins in the plant materials can be maintained to the largest extent.

Description

technical field [0001] The invention relates to a method for extracting total plant protein. Background technique [0002] Sample preparation is a key factor in proteomics research, and the quality of sample preparation almost directly determines the outcome of downstream analysis. An ideal sample preparation method should be able to dissolve all proteins to the greatest extent, while minimizing protein degradation, modification and non-protein contamination, and have good repeatability. [0003] Especially for plant tissues, which not only contain a large number of cell wall structures, proteases and oxidases, but also contain a large number of polysaccharides and secondary metabolites (such as lipids, phenols, starch, etc.), these unfavorable factors make it difficult to obtain Non-polluting, non-modified, complete and high-quality plant cell protein. Contents of the invention [0004] The purpose of the present invention is to provide a method for extracting total pla...

Claims

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Application Information

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IPC IPC(8): C07K1/30G01N1/28
Inventor 王绍辉杨瑞郝敬虹邢佳毅
Owner BEIJING UNIV OF AGRI
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