Method for realizing transmembrane transduction of marine microorganism low-temperature lipase gene by utilizing PTD-Tat

Abstract

The invention is applicable to the technical field of fusion expression of transmembrane transduction gene fragments and lipase, and provides a method for realizing transmembrane transduction of marine microorganism low-temperature lipase gene by utilizing PTD-Tat, the method comprises the following steps: 1, designing to obtain Lipase-tat fusion genes; 2, constructing a pET28a-Lipase-tat carrier: firstly, carrying out enzyme digestion on Lipase-tat by using Nco I and Xho I, and carrying out gel recovery on enzyme digestion fragments; then carrying out enzyme digestion on the pET28a carrier by using Nco I and Xho I, and carrying out gel recovery to obtain a linearized pET28a carrier fragment; connecting the enzyme digestion fragment with a pET28a vector fragment to obtain a recombinant expression vector plasmid; and 3, carrying out prokaryotic expression, purification and extraction on the Lipase-tat protein. Therefore, transmembrane transduction of the marine microorganism low-temperature lipase gene can be realized, so that the weight-reducing medicine prepared from the recombinant protein can enter cells; meanwhile, a theoretical and practical basis is provided for effective application of the transmembrane transduction mechanism of the PTD-Tat gene and the marine microorganism low-temperature lipase gene and protein drug delivery.

Description

technical field [0001] The invention relates to the technical field of fusion expression of transmembrane transduction gene fragments and lipase, in particular to a method for transmembrane transduction of low-temperature lipase gene of marine microorganisms by using PTD-Tat. Background technique [0002] With the development of society and economy, obesity, as a multifactorial chronic metabolic disease, is increasing day by day, and has reached a historical peak in the past 20 years. At present, the wave of weight loss is sweeping across the world. Nowadays, almost all weight loss drugs use lipase inhibitors. Their working principle is to reduce the absorption of fat in substances by the human body by inhibiting the decomposition and catalysis of lipase in the intestinal tract on substances that have entered the human body but have not yet formed human body fat. Control and treat obesity. However, lipase inhibitors can only work in the intestinal tract and cannot enter hum...

AI Technical Summary

Problems solved by technology

[0005] If PTD-Tat is used to carry the low-temperature lipase gene of marine microorganisms to achieve transmembrane transduction, it can not only decompose the substances that have entered the human body but have not yet formed human fat in the intestinal tract, but also enter the deep tissues of the human body for drug action , this way of losing weight will be a brand-new idea of ​​losing weight, but there is no research report on the method of using PTD-Tat for lipase transmembrane transduction

[0006] In summary, there are obviously inconveniences and defects in the actual use of the existing technology, so it is necessary to improve

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