Hybrid expression cassettes consisting of a milk protein promoter and a cytomegalovirus enhancer significantly increase mammary-specific expression of human lactoferrin in transgenic mice.
It is very important to develop an effective, specific, and robust expression cassette that ensures a high level of expression in the mammary glands. In this study, we designed and constructed a series of mammary gland-specific vectors containing a complex hybrid promoter/enhancer by utilizing promoter sequences from milk proteins (i.e., goat β-casein, bovine αs1-casein, or goat β-lactoglobulin) and cytomegalovirus enhancer sequences; vectors containing a single milk protein promoter served as controls. Chicken β-globin insulator sequences were also included in some of these vectors. The expression of constructs was analyzed through the generation of transgenic mice. Enzyme-linked immunosorbent assay (ELISA) analysis revealed that the hybrid promoter/enhancer could drive the expression of recombinant human lactoferrin (rhLF) cDNA at high levels (1.17-8.10 mg/ml) in the milk of transgenic mice, whereas control promoters achieved a very low rhLF expression (7-40 ng/ml). Moreover, the expression of rhLF was not detected in the serum or saliva of any transgenic animal. This result shows that all constructs, driven by the hybrid promoter/enhancer, had high mammary gland-specific expression pattern. Together, our results suggest that the use of a hybrid promoter/enhancer is a valuable alternative approach for increasing mammary-specific expression of recombinant hLF in a transgenic mouse model.
Engineering Research Centre for Transgenic Animal Pharmaceutics in Jiangsu Province, College of Veterinary Medicine, Yangzhou University, No. 12 Wenhui Rd., Yangzhou, Jiangsu, PR China. chengyong@yzu.edu.cn
Yong
Y
Cheng
Li-You
LY
An
Yu-Guo
YG
Yuan
Yi
Y
Wang
Fu-Liang
FL
Du
Bao-Li
BL
Yu
Zheng-Hong
ZH
Zhang
Yu-Zheng
YZ
Huang
Ting-Jia
TJ
Yang
D000818:Animals; D002417:Cattle; D002645:Chickens; D003587:Cytomegalovirus / Q000235:genetics*; D004742:Enhancer Elements, Genetic / Q000502:physiology*; D005260:Female; D006041:Goats; D006801:Humans; D038101:Insulator Elements / Q000235:genetics; D007781:Lactoferrin / Q000096:biosynthesis* / Q000235:genetics; D008321:Mammary Glands, Animal / Q000502:physiology*; D051379:Mice; D008822:Mice, Transgenic; D009928:Organ Specificity / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000502:physiology*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D055544:beta-Globins / Q000235:genetics
D011994:Recombinant Proteins; D055544:beta-Globins; D007781:Lactoferrin