Preliminary Studies On The Biosyntheses Of Heterogeneous Proteins In E.coli Cell-free System

The cell-free expression system is an in vitro translation system, compare with the in vivo translation system. Cell-free synthesis system enables the direct in vitro expression of protein from extrinsic template DNA or mRNA. It synthesizes protein through complementing substrate and energy in the cell extraction. In the new era of "bio-omics", the cell-free expression system opens a new strategy for protem expression, and is used widely because it can produce proteins in a mode of rapid, convenient and high-throughput expression.E. coli cell-free expression system is one widely used system which can synthesize protein effectively through complementing substrate and energy. The E. coli cell-free expression system was constructed and optimized in the present paper. The orthogal design method was used to improve the productivity in this E. coli cell-free system. Then six vectors were constructed for the purpose of expressing six HIV genes in E. coli cell-free system. Among them, VIF gene can be expressed effectively in this cell-free system. This work may be helpful to further expression of HIV genes in the cell-free system. Finally, two more genes (one SAM biosynthetic gene and one ALA biosynthetic gene) were inserted into cell-free vectors for the further expression of these two enzymes in the cell-free systems.