Establishing method of CD36 mutant gene stable eukaryotic expression cell line causing CD36 deletion

Abstract

The invention provides an establishing method of a CD36 mutant gene stable eukaryotic expression cell line causing CD36 deletion. The method comprises a while process of extracting total RNA from a CD36 deletion individual and conducting reverse transcription so as to obtain CD36 cDNA of whole segment of a CD36 mutant gene open reading frame; establishing a recombinant cloning vector containing the CD36 mutant gene cDNA and an EGFP fluorescent reporter gene, transforming the recombinant cloning vector into escherichia coli, and conducting plasmid amplification and identification; and transfecting the recombinant cloning vector into CHO-K1 cells, and conducting screening and identification so as to obtain the CD36 mutant gene stable eukaryotic expression cell line. The CD36 is distributed in a plurality of cells and tissues, and CD36 encoding gene mutation can cause the CD36 deletion in the individual. The CD36 takes an important effect in various physiological and pathological processes, involving such diseases as platelet homologous immunization, hypercholesteremia, diabetes, malaria and the like. With the application, the CD36 mutant gene stable eukaryotic expression cell line, which is established by virtue of the technology, can provide a resource and a working platform for researching the structure and the functions of the CD36, explaining physiological and pathological mechanisms which the CD36 participate in and discussing a role in clinical treatment and drug research and development.

Description

technical field [0001] The invention relates to biomedicine, in particular to a method for establishing a eukaryotic stable expression cell line of a CD36 mutant gene that causes human CD36 loss. Background technique [0002] CD36 (leukocyte differentiation antigen 36), also known as platelet glycoprotein IV (GPIV), GP88, GPIIIb, FAT or SCARB3, belongs to the class B scavenger receptor transmembrane glycoprotein family. The protein molecule can be extensively glycosylated with a molecular mass of about 88,000. The human CD36 protein consists of 472 amino acids. CD36 protein is widely distributed in a variety of cells and tissues in the human body, including platelets, monocytes / macrophages, microvascular endothelial cells, brain microglia, astrocytes, cardiac and skeletal muscles, adipocytes, tree dendritic cells, retinal epithelial cells, and breast and kidney tissues. Its main functions include acting as receptors for thrombin receptors, collagen receptors, long-chain f...

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Problems solved by technology

[0008] The currently reported researches on CD36 gene mutations that lead to CD36 deletion mostly focus on CD36 cDNA sequencing and prokaryotic cloning. There are no reports on the method of establishing eukaryotic stable expression cell lines of CD36 mutant genes that cause CD36 deletion in humans and related patents

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