Manufacturing method and application of insect in-vitro protein interaction detecting system

Abstract

The invention discloses a manufacturing method and application of an insect in-vitro protein interaction detecting system. At the positions of amino acid residues at the 159th bit and the 160th bit, mCherry is divided into a segment NmC and a segment CmC through a PCR method, and then the segment NmC and the segment CmC are each fused with a connecting zone sequence, a c-Myc label and a poly(A) sequence to form four fused segments; the four fused segments are each connected with a pIE-MCS plasmid with an autographa californica multiple nuclear polyhedrosis virus ie1 gene promoter and a gp64 gene poly(A) sequence to establish expression plasmids; the expression plasmids are combined to form an insect cell dimolecular fluorescent complementary detecting system. The method is simple and easy to implement, and the established dimolecular fluorescent complementary detecting system expands the application of an insect cell expression system and provides a convenient and effective technological means for insect proteomics and research on protein interaction between insects and pathogenic microorganisms of the insects.

Description

technical field [0001] The invention belongs to the technical field of bioengineering, and relates to a preparation method and application of an insect in vitro protein interaction detection system. Background technique [0002] Under the normal physiological metabolic conditions of cells, proteins are the executors of cell activity and function. Through complex interactions, proteins participate in various life processes such as cell signal transduction, proliferation, differentiation, immunity, and apoptosis. In recent years, with the acquisition of genome and transcriptome information of a large number of species, proteomics—the systematic study of the protein structure, function, and interaction network of a species has become a hot spot in the field of life sciences. Among them, analyzing protein interactions is the basis for revealing protein structure and function, which helps to systematically explain various life phenomena and processes, explore the occurrence and ...

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Problems solved by technology

[0010] The purpose of the present invention is to provide a convenient and effective in vitro protein interaction detection system for insects, aiming to solve the problem that the background of the existing bimolecular fluorescence complementary detection method is relatively high, and the promoter system cannot be recognized and guided in insect cells Target protein expression, the detection of protein interactions between insects and their pathogenic microorganisms cannot be performed using this type of system, the efficiency of BIFC plasmid transfection in mammalian cells is low, it is not suitable for large-scale protein interaction screening and detection, and the process of mammalian cell culture is complicated, etc. question

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