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Method for identifying glutenin macro-polymer content in wheat
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A technology for gluten and macromers is applied in the field of identifying wheat gluten macromers, which can solve the problems of poor particle size classification ability, low accuracy, and inability to effectively separate gluten aggregates.
Inactive Publication Date: 2015-03-25
CAPITAL NORMAL UNIVERSITY
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Since the molecular weight of glutenin macromers can reach millions of Daltons, the commonly used polyacrylamidegel electrophoresis (SDS-PAGE) cannot effectively separate them. Therefore, multi-layer stacking gel SDS-PAGE (MS-SDS-PAGE) is currently used. PAGE), this method is based on SDS-PAGE, the advantage is that different gel concentration gradients can be designed, and the particle size distribution of glutenin macromers in gels with different concentrations can be compared by means of a gel scanning system. Second, at present, the biuret colorimetric method is mostly used in China. The advantage is that it is simple to operate and requires simple equipment. The disadvantage is that the accuracy is low and the ability to classify the particle size of glutenin aggregates is poor.
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Embodiment 1
[0057] 1. Material sampling
[0058] The seeds of wheat variety Zhongyou 9507 were used as test materials, the seeds were randomly selected, and 20-30 mg was weighed after grinding for the extraction of glutenin macromers and gel exclusion high performance liquid chromatography analysis.
[0060] Extraction method of glutenin macropolymer (GMP):
[0061] 1) Grind the wheat seeds with liquid nitrogen, weigh 20 mg powder and put it into a 2.0ml centrifuge tube, add 1.8ml 0.05M phosphate buffer solution containing 2% SDS, vortex for 30min, centrifuge at 13000rpm for 10min, remove the supernatant, for the EPP;
[0062] 2) Add 1.8ml of 0.05M phosphate buffer containing 2% SDS to the precipitate again, and vortex to fully mix the precipitate with the buffer;
[0063] 3) Use an ultrasonic cell disruptor to ultrasonically disrupt the precipitate, with the ultrasonic set at 200W, 30s, 5 times.
[0064] 4) Vortex the sonicated suspension for 2 hours; ...
Embodiment 2
[0092] 1. Material sampling
[0093] The seeds of wheat varieties Imbross, Jing 411, Zhongguochun and Jingdong 8 were used as test materials respectively, and 20-30 mg of seeds were randomly selected after being ground and weighed for gluten extraction and ultra-high performance liquid chromatography analysis.
[0096] 1) Grind the wheat seeds with liquid nitrogen, weigh 20 mg powder and put it into a 2.0ml centrifuge tube, add 1.8ml 0.05M phosphate buffer solution containing 2% SDS, vortex for 30min, centrifuge at 13000rpm for 10min, remove the supernatant, for the EPP;
[0097] 2) Add 1.8ml of 0.05M phosphate buffer containing 2% SDS to the precipitate again, and vortex to fully mix the precipitate with the buffer;
[0098] 3) Use an ultrasonic cell disruptor to ultrasonically disrupt the precipitate, with the ultrasonic set at 200W, 30s, 5 times.
[0099] 4) Vortex the sonicated suspe...
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Abstract
The invention discloses a method for identifying a gluteninmacro-polymer in wheat. The method comprises the following steps: S1: providing wheat grains and extracting a gluteninmacro-polymer in the wheat grains by virtue of a buffer solution to obtain a solution; S2: carrying out ultrasonic treatment on the suspension so that partial gluteninmacro-polymer is broken and is dissolved in the buffer solution; S3: filtering the buffer solution with the dissolved glutenin macro-polymer by virtue of an aqueous filter membrane so as to obtain a to-be-detected sample; and S4: carrying out size exclusion chromatography analysis on the to-be-detected sample, and implementing automatic integration to obtain the expression quantity of the glutenin macro-polymer in the wheat grains. According to the embodiment disclosed by the invention, the method can be used for rapidly and accurately identifying the content of the glutenin macro-polymer in the wheat grains in the different stages of grain development, and can be used for effectively screening the varieties of the glutenin macro-polymer according to different expression quantities in mature grains.
Description
technical field [0001] The invention relates to the technical field of identifying wheat glutenin macromers, more specifically, relates to a method for identifying the content of wheat glutenin macromers through gel chromatography (SE-HPLC) technology. Background technique [0002] Wheat grains are rich in starch, more protein and a small amount of fat, and are an important source of plantprotein for humans. In my country, wheat is mainly used for flour processing, and the processing quality of wheat is mainly affected by factors such as growth environment, agronomic measures and intrinsic genotype, which determine the content and quality of wheat protein. According to the different solubility of wheat grain protein in water, NaCl solution, ethanol and dilute acid or dilute alkali solution, wheat grain protein is divided into four categories, namely water-soluble protein soluble in water, salt-soluble protein soluble in salt solutionProtein, gliadin soluble in alcohol and...
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