The Studies Of Lentivirus Carrier Sperm Mediated Method Produce Transgenic Chicken

In this study,we explore and study the efficient way of production based on the lentiviral carrier sperm mediated method to establish transgenic technology system of chicken.This study build eukaryotic expression vector and lentiviral expression vector with green fluorescent protein gene, contrast cell phenotype change and transfection efficiency with two expression vectors transfecting293T cells. According to the Index of viral infection,we infect the primary cultured cells of chicken primordial germ cells and intestine epithelial cells with lentivirus,and test cells infection with real-time PCR.We observed and analyzed the rooster reproductive organs organizational structure with mature cock testicular tissue sections,and injected the lentiviral a green fluorescent protein reporter gene into mature rooster testicles using minimally invasive surgical techniques.The sperm expression of the green fluorescence were collectd to observe.The results:We construct eukaryotic expression vector PcDNA3.1-GFP and lentiviral expression vector that reporter green fluorescent protein gene and expressed in293T cells.The number of GFP cells is similar between two vectors transfacted cells.The cells transfected by eukaryotic expression vector PcDNA3.1-GFP evenly distributed,still monolayed.The cell infected by Lentivirus is in colonies distribution obvious phenotypic changes with strong fluorescence signal.The Real-time quantitative PCR results of cells transfected by Lentivirus:the multiplicity of infection in293T cells is6.08×107TU/mL,in chicken embryo intestinal epithelial cells is5.36×105TU/mL,in chicken embryo primordial germ cells is0.84×102TU/mL.Mature cock testicular tissue sections shows that cock testicular parenchyma composit the seminiferous tubules mesh, not divided into lobules and suitable for testicular injection.