Produce Transgenic Mice By Direct Injection EGFP-liposome Plasmid To Testis And Ovaries In Vivo

Reproductive cells can transmit parental genetic information to the offspring, making them an attractive target cell population for animal transgensis. In this paper, we describe a novel approach for producing transgenic mice by transfected reproductive cells in vivo, and the female mice produced transgenic offspring after mating with transgenic males.The experiment use enhanced green fluorescent protein (eGFP) as a marker. The eGFP plasmid was cultivated with liposome then injected into testicle and ovaries of Kunming white mice. Different periods male and female reproductive cells was transfection, therefore the offspring expressing eGFP was obtained. The length of the plasmid which contains green fluorescent protein gene is 5.2kb. Before injection, the cDNA was extracted, purified and identificated. Plasmid -Liposome Complex was injected into testicle and ovaries inorder to study new methods of transgenic animals. All the future generations of mice were divided into three groups. The A group is given the birth by transgenic male mouse and wild-tape female mouse. The B is by transgenic female mouse and wild-tape male mouse. And the parents of the mice of C are transgenic mice.It's confirmed that the reproductive capacity of male mice whose testis was injected in one-side and two-side was no significant difference. Foreign gene had been gradually degradation after being injected into the testes. The best time of using the mice is in after surgery 40days. The time of degradation of foreign gene in ovary of female mice is 5 days.The 2-cell embryo which was obtained by transgenic female mice mating with wide-tape males expresses eGFP.The fequency of transgenic mouse production was determined by pair-mating with the 3 group of mice, and by polymerase chain reaction (PCR) and sequence of DNA taken from the tails of the offspring. In the A group, among the 79 F1 transgenic mice, 32mice were transgenic mice. The transgenic efficiency was 40.51%.The B is 48.94%(46/94),and C is 81.36% (48/59) .The expression of eGFP gene in the transgenic mice was detected by RT-PCR and vivi-formatter of green fluorescent. The results showed that the introduxed gene was detectable in the gonads/lung/kidney/tail of the F1 mice.The results demonstrate that the foreign gene and liposome derect inject into testis and ovary can be used to successfully generate transgenic mice. The founder mice reansmitted the eGFP to their offspring. Additionally, the reporter gene was not only integrated into the genome, but was also transcribed, translated into a functional protein, and reansferred to the next generation. Thus, oocytes and spernid transferred at the same time is efficiency to abtain the transgenic offspring. The technique overcomes the drawback of the in vitro transduction approach, and will be useful as a novel method for producing transgenic animals.