Recombinant human collagen and its coding gene and preparation method thereof

Abstract

The invention provides recombinant human collagen and a method for preparing the recombinant human collagen. The method comprises the following steps: 1)preparing plasmid: performing artificial whole gene synthesis of a sequence of SEQ ID No:3 in a sequence table, then performing Xho I and EcoR I double enzyme digestion on a pPIC9K carrier and human collagen in SEQ ID No:3 through artificial whole gene synthesis, then extracting plasmid; 2) converting: mixing the prepared plasmid and pichia yeast for conversion to obtained converted bacterium colony; 3) screening of multicopy additive recombinants; 4) obtaining a broth through fermentation; and 5) purifying to obtain the product. The invention also provides a gene sequence for coding the recombinant human collagen. The recombinant human collagen has good biological compatibility, and the purity can reach more than 95%.

Description

technical field [0001] The invention belongs to the field of bioengineering, and specifically relates to a gene recombinant human collagen and its coding gene, and also relates to a preparation method of the recombinant human collagen. Background technique [0002] Collagen is the most abundant protein in the body, accounting for about 25%-33% of the total protein. It is widely present in the bones, tendons, cartilage, skin and other connective tissues of the human body. It is an extracellular matrix ( ECM) plays an important role in maintaining the normal physiological functions and repairing of cells, tissues and organs. In terms of molecular structure, collagen is composed of parallel linear chains, and each linear chain is composed of three twisted left-handed α-peptide chains tightly combined through inter-chain interactions to form a very strong right-handed triple helix structure. Each α-peptide chain consists of more than 300 repeated Gly-X-Y triplets, and the two e...

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Problems solved by technology

Some research institutions and biological companies at home and abroad have successively invested in the research and development of recombinant collagen. In 2012, Yang Xia et al. achieved high expression in E. coli by repeating type III collagen peptides and fusing them with type II collagen peptides; Fan Daidi from Northwest University in my country used Escherichia coli to produce human-like collagen through high-density fermentation culture, obtained highly expressed human-like collagen, and used it as raw material to develop tissue engineering materials and cosmetics, but part of the amino acid sequence they designed was not human collagen sequence, and at the same time , the pyrogens produced by bacterial expression make the expression product difficult to apply clinically, and the target protein is often expressed in the form of inclusion bodies, which makes product purification difficult. In addition, the post-translational modification processing system of the prokaryotic expression system is imperfect, and the biological activity of the expression product is low. and other defects affect the quality of the product

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