Purity identification method of millet hybrid based on single-grain prolamin extraction and detection technology

Abstract

The invention discloses a method for identifying the purity of millet hybrids based on single-grain prolamin extraction and detection technology, which improves the solubility and extraction of gliadin in single-grain millet seeds by adopting a multi-solvent compounded gliadin extraction solution. Efficiency; by increasing the gel concentration to more than 15% to achieve sufficient interception of prolamin, by increasing the amount of ferrous sulfate and hydrogen peroxide to increase the speed and strength of the redox reaction to increase the gel strength and reduce the process of the glue on the board The damage rate in the middle, makes the gliadin band spectrum fully and clearly displayed; finally, the band spectrum is read to complete the identification of the purity of millet hybrids.

Description

technical field [0001] The invention relates to the technical field of detection and identification of biomolecules, in particular to a method for extracting and detecting prolamin from single grain millet seeds and its application. Background technique [0002] In recent years, great achievements have been made in the utilization of millet heterosis. For example, "Zhangzagu No. 5" bred by Zhangjiakou Academy of Agricultural Sciences in 2007 created a high-yield record of 810.6 kg per mu in a small area of ​​spring valley in my country, and was selected by Tangshan Normal University. Yu's "5695" created a high-yield record of 765.5 kilograms per mu in a small area. However, since millet hybrids are two-line hybrids, that is, there are only female parent-sterile lines and male parent-restorer lines, there is no maintainer line, and the female parent sterile line is not 100% sterile, but has about 5% sterility. Self-seeding characteristics, and can be stably inherited, the mai...

AI Technical Summary

Problems solved by technology

[0003] Before the 1990s, the identification of millet hybrids was mainly based on the difference in the seedling color of the parents. The material with darker seedling color was selected as the male parent, and the male sterile line with lighter seedling color was used as the female parent. According to the genetic law of sex, the color of the hybrid is the same as that of the male parent. By removing the female parent with a lighter color mixed in the hybrid, the hybrid is removed, and the purity of the hybrid is determined by investigating the proportion of the sterile plants with a lighter color. Although this method can remove some hybrids in hybrids, it limits the application of darker sterile lines, and is labor-intensive and time-consuming. It cannot remove mixed restorer lines and other materials in hybrids, and the purity of identification is not accurate.

[0004] After the 1990s, with the introduction and successful creation of herbicide-resistant materials, through the cultivation of herbicide-resistant restorer lines, the sterile lines are not herbicide-resistant, and the herbicide-resistant traits are dominantly inherited. Spray herbicides to kill the false hybrids among them to identify the purity of the hybrids and remove the false hybrids. This method makes it possible to apply CMS lines of various seedling colors, but it is still unable to identify the recovery of mixed hybrids. Lines or other herbicide-resistant materials, still can not completely remove the false hybrids in the hybrids, and can not accurately identify the purity of the hybrids

[0008] Through research, some millet researchers have concluded that salt-soluble protein can be used for variety identification [13] , but further studies have shown that most of the non-prolamins such as salt-soluble proteins are non-storage proteins that participate in the immune response, and their polymorphism is poor, which is not suitable for the identification of variety purity. [14] , on this basis, the researchers began to try to use the wheat prolamin A-PAGE method to identify polymorphisms of glutinous prolamin [5,14,15,16] , but due to the small grain size of millet, more than 10 seeds are needed to extract prolamin from millet [14] , and the variety purity identification must be based on each seed, therefore, this method cannot be used for the purity identification of millet hybrids; researchers have begun to use the SDS-PAGE method to identify polymorphisms of millet prolamins [17] However, due to the shortcoming of long electrophoresis time in SDS-PAGE, and when the concentration of isopropanol is increased to 50%, more than 20 grains of prolamin are needed to extract prolamin and detect polymorphism , can not meet the needs of the identification of the purity of millet hybrids

So far, the prolamin polymorphisms of glutinous rice cannot be applied to the identification of genetic diversity and variety purity

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