Identification method of highly stoichiometric non-germline mutation in eucaryotic cell

Abstract

The invention relates to an identification method of the high stoichiometric ratio non-germline mutation in the eucaryotic cell and provides an semi-quantitative identification method of the protein synthesis error caused by the single amino acid mutation of the non-germline mutation (NGM), wherein the method is based on the proteomics technology and genome DNA sequencing of the mass spectrometry. The method is particularly as follows: the large-scale proteome analysis, the unrestricted sequence alignment and the identification and gene sequencing of the high resolution second-order mass spectrum and third-order mass spectrum of the synthetic peptide segment are integrated to perform systematic synthesis to the NGM and identify that the four peptide segments of four proteins contain the NGMs. By adopting the method for estimation, the NGM occurrence in the peptide segment is about 0.05%, which is more than 0.3% on the protein level. According to the peptide mass spectrum count ratio of mutant peptide segments to normal peptide segments, the stoichiometric ratio of the NGM is 0.05 to 0.71. By adopting the identification method, the NGM is proved to have high occurrence and stoichiometric ratio while the NGM is predicted to be used as a possible mechanism to join the basic biological and physiological processes.

Description

technical field [0001] The present invention relates to a method for identifying non-hereditary mutations, especially the application of the overall proteomics method combined with DNA sequencing and the use of affinity reagents that specifically bind to protein mutation residues and flanking sequences to identify high chemical mutations in eukaryotic cells. A method of quantifying non-inherited mutations. Background technique [0002] Genetic mutations are associated with diseases such as cancer, neurological diseases and metabolic abnormalities (1，2) . For cancer, one of the leading causes of death in Western countries, gene mutations are the main cause. Mutations in even 1% of genes in both somatic and germ cells can lead to cancer (3) . Thus, the study of the association of random genetic mutations with cancer has been a major area of ​​study in tumor biology over the past few decades, demonstrating the critical role of genome integrity in health and disease. [000...

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Problems solved by technology

However, given the typically high error rates of unrestricted sequence alignments, and the lack of stringent verification methods such as MS / MS for synthetic peptides, the reliability of the above studies has not yet been fully assured

Furthermore, the fundamental question of whether mutations occur in the DNA sequence or during protein synthesis remains unanswered

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