Proteome identification method and applications thereof

Abstract

The present invention discloses a proteome identification method and applications thereof. The proteome identification method comprises: 1) extracting the total protein of an organism, and dissolving the total protein by using a protein lysate to obtain a protein solution; 2) sequentially carrying out reducing and alkylation on the protein in the protein solution to obtain a reduced alkylated product; 3) carrying out enzymolysis on the protein in the reduced alkylated product by using a protease to obtain an enzymolysis product; 4) labeling the protein in the enzymolysis product by using an iTRAQ reagent to obtain a labeled product; 5) pre-grading the labeled product by using UPLC-Q-TOF-MS to obtain different small proteome samples, carrying out primary identification on each small proteome samples by using MALDI-TOF-TOF-MS, and combining the group having the absorption peak being not covered to obtain different large proteome samples; and 6) carrying out proteome identification on different large groups of the samples by using NanoLC-Q-TOF-MS to obtain the proteome identification data.

Description

technical field [0001] The invention relates to a proteome identification method and its application in the field of biotechnology. Background technique [0002] Protein is the product of gene expression, and it is also the main executor and embodiment of gene function. The study of protein is of great significance to reveal the function of gene and the essence of life phenomenon. Proteomics is an emerging discipline that emerged under the background of breakthroughs in genomics research and high-throughput protein analysis technology. research to elucidate its biological function. Currently, proteomics technologies include two-dimensional gel electrophoresis, two-dimensional fluorescent differential gel electrophoresis, isotope affinity labeling technology, and isotope-labeled protein quantitative technology, including various protein separation and identification technologies. [0003] Isotopic tags for relative and absolute quantitation (iTRAQ) technology is a new and p...

AI Technical Summary

Problems solved by technology

During iTRAQ's nano-liter mass spectrometry identification, high-abundance glutenin will cover the mass spectrometry signals of low-abundance albumin and globulin, thereby reducing the number of iTRAQ identifications

In addition, due to the incomplete sequencing of the whole wheat genome, the number of wheat protein entries in the database is far less than that of Arabidopsis, rice and other species, which also increases the difficulty of mass spectrometry identification of wheat proteins

[0008] At present, there are few reports on the identification of wheat seed proteome using iTRAQ technology. Ma et al. (2014) used iTRAQ tandem LC-MS / MS technology for the first time to analyze the protein expression profile of wheat seed development. They identified 1146 proteins in total. The number of proteins is far lower than the number of proteins predicted by the genome. Therefore, improving the extraction method of wheat seed protein components and increasing the number of proteins identified by iTRAQ mass spectrometry are of great significance and substantial promotion for the study of wheat seed proteome.

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