Construction Of Sheep Beta-Defensin-1 Gene Eukaryotic Expression Vector And Expression In Bovine Fetal Fibroblast Cells

β-defensin is an important members of the family and is mainly expressed in the skin, mucous membranes and other epithelial tissue of vertebrate, constitute the body against microbial invasion of the first chemical barrier to mucosal surfaces has been recognized as an integral anti-microbial barrier components.In order to preparation A lot of antibacterial activity, low toxicity ofβ-defensin by biological engineering technology, the recombinant plasmid PMD19- TSinple- sBD-1 which was preserved in our laboratory with target sBD-1 gene was amplified by specific primers. The purified PCR product was cloned in a eukaryotic expression vector pEGFP-C1 with the enhanced green fluorescent protein. The recombined eukaryotic expression vector, pEGFP-C1-sBD-1, was transfected into bovine fetal fibroblast cells mediated by Liposome.The expression of sBD-1 in bovine fetal fibroblast cells was examined by real time RT-PCRThe result shows that the target sBD-1 gene was obtained. After the PCR analysis of recombinant plasmid and the result of cDNA sequencing demonstrated that with the homology is 100%.These results show that the vector completely correct and can be transfected. After 24h, we observed by fluorescence microscope with a large number of green fluorescence, proved sBD-1 gene can be expressed in eukaryotic cells. Application of real-time fluorescence quantitative PCR detection showed that sBD-1 gene in the bovine fetal fibroblast cells can be highly expressed. To further study the sheepβ-defensin function and the vector of animal mammary gland bioreactor used in the research foundation. To further study the sheepβ-defensin function and the vector of animal mammary gland bioreactor used in the research foundation.