Non-degeneration plant holoprotein extracting solution and preparing method thereof

A technology of extracting liquid and whole protein, which is applied in the field of plant physiology and biochemistry, and can solve the problems of inability to analyze enzyme activity, protein denaturation, and inability to accurately quantitatively compare enzyme activity.

Active Publication Date: 2016-01-13
ZHENGZHOU TOBACCO RES INST OF CNTC
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  • Abstract
  • Description
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Problems solved by technology

Urea denatures proteins, making it impossible to analyze enzyme activity; while reducing agents such as DTT seriously interfere with BCA, Bradford, and other methods for measuring protein concentration, making accurate quantitative comparisons of enzyme activities impossible
When many laboratories extract whole plant protein for physiological and biochemical analysis, in order to avoid protein denaturation and degradation, it is necessary to analyze it immediately after extraction, which is obviously unrealistic for large quantities of samples

Method used

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  • Non-degeneration plant holoprotein extracting solution and preparing method thereof

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Embodiment Construction

[0038] The present invention is described in further detail below by a specific embodiment:

[0039] 1. Prepare the extract

[0040] Prepare 1L extract: containing 100mM phosphate buffer, 5mM EDTA, 5% glycerol, 2.5% PVP and protease inhibitors:

[0041] Weigh 29.367 grams of Na 2 HPO 4 ?12H 2 O and 2.808 g NaH 2 PO 4 ?2H 2 O was added to a 1L beaker and dissolved as a phosphate buffer solution with pH 7.5 and 100mM;

[0042] Add 25ml of 200mM EDTA-Na mother solution with pH 7.5;

[0043] Place the beaker directly on the percent scale and weigh 50 grams of glycerin;

[0044] Add a stir bar and stir well with a magnetic stirrer;

[0045] Use a graduated cylinder to add deionized water to make up to 1L;

[0046] Pour into a 1L reagent bottle, loosely screw on the bottle cap, fix the bottle cap to the bottle body with a sterilization indicator strip, and sterilize at high temperature;

[0047] After cooling to room temperature, tighten the cap and store at 4°C for later...

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Abstract

The invention relates to plant protein extracting, in particular to a non-degeneration plant holoprotein extracting solution and a preparing method thereof. The extracting solution is characterized by being an aqueous solution containing a phosphate buffer solution, EDTA, glycerol, PVP and protease inhibitors. The extracting solution has the advantages that plant holoprotein extracted with the holoprotein extracting solution is suitable for analyzing the activity of protease and measuring the protein concentration with methods such as BCA; the holoprotein is not degenerated, reducing agents influencing measuring of the BCA and the like are not added, the extracted holoprotein can be stored for several months in the frozen mode, and sediment is basically avoided after re-melting; in addition, the activity of the holoprotein is basically free of influences before and after storage, the extracted holoprotein is high in amount and can be further directly used for further separation purification, and therefore the extracting solution has important application value in the aspect of plant physiology and biochemistry researching.

Description

technical field [0001] The invention relates to plant protein extraction and belongs to the technical field of plant physiology and biochemistry, in particular to a non-denatured plant whole protein extract and a preparation method thereof. Background technique [0002] The study of plant physiology and biochemistry is inseparable from the detection of related enzyme activities, such as the changes of SOD enzyme activity and GST enzyme activity that are often concerned under different stress conditions. Due to the specificity of different enzyme activities, the use of whole protein extracts for detection does not require purification of the target protein, which is time-saving, labor-saving, and reliable. At present, there are many plant protein extracts on the market, which are generally used in subsequent proteomics research, and are characterized by the addition of urea, DTT, etc. Urea denatures proteins, making enzyme activity analysis impossible; and reducing agents su...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C07K1/14
Inventor 周会娜刘萍萍翟妞陈霞陈千思毛李鸿郑庆霞徐国云张慧林福呈
Owner ZHENGZHOU TOBACCO RES INST OF CNTC
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