EGFP转基因大鼠的建立

Establishment of EGFP Transgenic Rat

目的建立增强绿色荧光蛋白、（EGFP）转基因大鼠，为器官移植、基因治疗提供理想的动物模型。方法构建带有CMV—EGFP的重组慢病毒；通过大鼠受精卵透明带下显微注射的方法感染受精卵，并将注射后的受精卵移植到假孕母鼠输卵管中，出生的仔鼠通过PCR方法鉴定，获得首建大鼠；并通过RealTimePCR、Western—blot和荧光观察进行表达分析。结果显微注射后，210枚成活的受精卵移植到假孕母鼠；经繁育，出生2只大鼠；经PCR鉴定，有1只为阳性，阳性率为50％；经RNA、蛋白水平表达的检测及荧光观察，该大鼠表皮、骨骼肌、胰腺、肾脏、心脏、睾丸等均有EGFP表达。结论初步建立了转基因大鼠技术，成功获得了可稳定表达EGFP的转基因荧光大鼠。

Objective To establish an enhanced green fluorescent protein （EGFP） transgenic rat model, which will be used for organ transplant and gene therapy. Methods The lentiviruses carrying the EGFP driven by the CMV （cytomegalovirus） promoter were generated and injected into the perivi- telline spaceof single-cell rat embryos obtained from superovulated Sprague Dawley females mated on the previous evening with Sprague Dawley males. The injected zygotes were transferred into oviduct of pseudopregnant SD females. Transgenic rat were identified by PCR analysis on genomic DNA from rat tail, the transcription and translation pattern of EGFP gene were detected by Real Time-PCR, Western- blot and fluorescent observation. Results After microinjection 210 injected zygotes were transferred. A line of EGFP transgenic rat was established. The green fluorescent protein was expressed in most of the tissues from the transgenic rat line, such as skin, muscle, pancreas, kidney, heart, testis, and so on. Conclusion The transgenic rat technical platform is established.