Yan Hua, Li Guo-cai, Sun Huai-chang
Department of Pathogenic Biology and Immunology, School of Medicine, Yangzhou University, Yangzhou, Jiangsu, PR China.
Zhonghua Yi Xue Yi Chuan Xue Za Zhi. 2005 Oct;22(5):541-4.
To evaluate the feasibility of generating animal mammary gland bioreactors expressing human lysozyme (hLYZ).
The recombinant vector p205C3-hLYZ, as a result of connecting the hLYZ cDNA with the mammry gland expression vector p205C3, was used to generate transfer genic mice by microinjection.
A total of 136 F0 mice were obtained, of which 7 (2 females and 5 males) and 4 (1 females and 3 males) were found to contain the transfer-gene by PCR and Southern blotting respectively. The results of Western blotting indicated that the expressed protein had the same molecular weight as that of normal hLYZ. From the F1 generation on, the mice mated only with their brothers or sisters and a colony of F7 transgenic mice was obtained. Among the offspring, the female transgenic mice maintained and expressed the transfer-gene stably with an expression level as high as 750 mg/L. The expressed protein had strong tissue specificity, and in addition to the mammary glands, some degree of ectropic expression in the spleens and intestines of the transgenic mice was confirmed by dot blotting assay.
These data indicate that the mice mammary gland bioreactors expressing hLYZ have been successfully generated.
评估生成表达人溶菌酶(hLYZ)的动物乳腺生物反应器的可行性。
将hLYZ cDNA与乳腺表达载体p205C3连接构建而成的重组载体p205C3 - hLYZ,通过显微注射用于制备转基因小鼠。
共获得136只F0小鼠,其中分别通过PCR和Southern印迹法检测发现7只(2只雌性和5只雄性)和4只(1只雌性和3只雄性)含有转基因。Western印迹结果表明,表达的蛋白与正常hLYZ具有相同的分子量。从F1代开始,小鼠仅与它们的兄弟姐妹交配,获得了一群F7转基因小鼠。在后代中,雌性转基因小鼠稳定维持并表达转基因,表达水平高达750 mg/L。表达的蛋白具有很强的组织特异性,除乳腺外,通过斑点印迹分析证实转基因小鼠的脾脏和肠道存在一定程度的异位表达。
这些数据表明已成功生成表达hLYZ的小鼠乳腺生物反应器。