Quick clone and expression plasmid vector containing N utilization substance A (NusA) protein fusion label

Abstract

The invention relates to a clone and expression plasmid vector. The clone and expression plasmid vector contains a green fluorescent protein (GFP) gene segment which contains a tobacco etch virus (TEV) enzyme recognition site and is used for encoding the gene sequence of NusA protein, and of which two sides are provided with bovine serum albumin (BsaI) restriction enzyme cutting sites. The constructed vector can realize the quick and efficient clone and protein expression of the targeted gene without depending on ligase. The NusA protein label and the TEV enzyme recognition site are introduced into the vector; and by the NusA protein label, the solubility of the insoluble protein can be improved, and the protein can be separated and purified conveniently; and a TEV enzyme recognition sequence is positioned at the downstream of the gene sequence of the NusA protein label, the purified protein is cut by TEV enzyme, and the NusA protein label can be cut off, so that the natural targeted protein can be obtained. The TEV enzyme is the enzyme which is the most economic and applicable at present, so that the experimental cost is greatly saved.

Description

technical field [0001] The invention belongs to the technical field of molecular biology, and in particular relates to a cloning and protein expression plasmid carrier, that is, a rapid cloning and protein expression plasmid carrier containing a NusA protein fusion tag. Background technique [0002] The traditional cloning method is to generate sticky ends of 2-4 bases by digesting the plasmid vector and the target gene fragment respectively with the same double enzyme digestion, and then use ligase to connect the complementary sticky ends, and then transform Escherichia coli cells, Express purified protein. Due to the relatively short length of the cohesive end, T4 DNA ligase must be used to ligate the vector and the target gene fragment, and some vectors may self-ligate during this process. This method takes about two to three days to clone a gene. In order to realize a ligase-independent cloning method, it is necessary to construct a more convenient and effective clonin...

AI Technical Summary

Problems solved by technology

Moreover, the commercial carriers currently on the market can use very limited fusion tags, and the protease designed on the carrier to separate the target protein from the tag is also relatively expensive.

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