title: A mWAP-hLF hybrid gene locus gave extremely high level expression of human lactoferrin in the milk of transgenic mice.
abstract: The production of pharmaceuticals by current mammary gland bioreactor techniques is limited by the low expression level of foreign proteins. We describe here a novel method that solves this problem. A successive three-step gap-repair strategy was developed to replace the genomic coding sequence in mouse whey acidic protein (mWAP) gene locus with that of human lactoferrin (hLF) precisely from the start code to the end code. A 50-kb mWAP-hLF hybrid gene locus was constructed, and corresponding transgenic mice were generated. An extremely high-level expression of rhLF in the milk was demonstrated by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and Western blot. The expression level ranged from 16.7 to 29.8 g/l among five transgenic lines, as indicated by the ELISA assay. Importantly, the expressed rhLF maintained the same antibacterial activity as the native hLF. Our strategy can very likely also be used for the efficient expression of other valuable pharmaceutical proteins.
authors.affiliation: Cell Engineering Department, Beijing Institute of Biotechnology, No. 20 Dongdajie Street, 100071, Beijing, China.
authors.forename: Gengshou
authors.initials: G
authors.lastname: Shi
authors.forename: Hongxing
authors.initials: H
authors.lastname: Chen
authors.forename: Xiaojie
authors.initials: X
authors.lastname: Wu
authors.forename: Yanrong
authors.initials: Y
authors.lastname: Zhou
authors.forename: Zhuguo
authors.initials: Z
authors.lastname: Liu
authors.forename: Tao
authors.initials: T
authors.lastname: Zheng
authors.forename: Peitang
authors.initials: P
authors.lastname: Huang
mesh_terms: D000818:Animals; D019149:Bioreactors; D002678:Chimera; D005260:Female; D018628:Gene Dosage; D005818:Genetic Engineering / Q000379:methods*; D005822:Genetic Vectors / Q000235:genetics*; D006801:Humans; D007781:Lactoferrin / Q000096:biosynthesis* / Q000235:genetics; D008321:Mammary Glands, Animal / Q000378:metabolism*; D051379:Mice; D008822:Mice, Transgenic; D008892:Milk / Q000378:metabolism*; D008894:Milk Proteins / Q000235:genetics; D014158:Transcription, Genetic
chemical_list: D008894:Milk Proteins; C024471:whey acidic proteins; D007781:Lactoferrin