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目的以携带绿色荧光蛋白(green fluorescent protein,GFP)的慢病毒为载体,制备转基因小鼠,建立慢病毒介导的转基因动物制备技术平台。方法慢病毒表达载体FUGW与ViraPowerTM Lentiviral Packaging Mix(1∶2)混合,用LipofectamineTM 2000转染293FT细胞。包装出的病毒经浓缩后以293T细胞测定病毒滴度。采用受精卵卵周隙显微注射的方式制备转基因小鼠。出生小鼠以PCR及荧光体视显微镜下镜检的方式进行鉴定。结果包装出的慢病毒经浓缩后滴度可达108IU/ml。F0代小鼠GFP PCR阳性率为70.27%(26/37)。荧光体视显微镜下观察荧光小鼠所占比例F0代为65.22%(15/23),F1代为55%(11/20),荧光强度两代个体相当。结论制备出携带GFP的可传代的转基因小鼠,建立了慢病毒载体介导的转基因小鼠制备技术平台。
OBJECTIVE: To construct transgenic mice with lentivirus carrying green fluorescent protein (GFP) as carrier, and to establish a lentivirus-mediated transgenic animal preparation technology platform. Methods The lentivirus expression vector FUGW was mixed with ViraPowerTM Lentiviral Packaging Mix (1: 2) and 293FT cells were transfected with LipofectamineTM 2000. The packaged virus was concentrated to 293T cells to determine the virus titer. Transgenic mice were prepared by microinjection of the zygote of zygotes. Born mice were identified by PCR and fluoroscopy microscopy. The results of packaging lentiviruses after concentration of up to 108IU / ml. The F0 generation mouse GFP PCR positive rate was 70.27% (26/37). Fluorescent microscopic observation of fluorescent mice accounted for the proportion of F0 on behalf of 65.22% (15/23), F1 generation was 55% (11/20), the fluorescence intensity of two generations of individuals quite. Conclusion GFP transfected transgenic mice were prepared and a lentiviral vector-mediated transgenic mouse preparation technology platform was established.