title: Investigation of hFVIII production in mammary glands of transgenic mice.
abstract: Hemophilia A is an X-linked disorder affecting 1 in 10,000 males. The disease is caused by a defect or mutation of factor 8 or 9. Human factor 8 gene (hFVIII) is a relatively large gene consisting of 26 exons and approximately 2,351 amino acids with a length of 9 Kb mRNA. Expression of hFVIII in mammalian milk is becoming a widespread strategy for high-level production of hFVIII because of the most complex post-translational modifications. The aim of this study was the cloning and expression of hFVIII in mammary glands of two transgenic mice. To obtain a recombinant plasmid, first a plasmid carrying an FVIII gene fragment (pCMV6-hFVIII) was digested by EcoRI-SalI restriction enzymes and then the fragment was purified from agarose gel and inserted into a pUCWAP7 vector carrying a tissue-specific promoter (mWAP 4.1 kbp). After that, it was isolated by agarose gel and transferred into the murine zygotes by standard microinjection methods. Methods for expression of recombinant FVIII RT-PCR and ELISA were studied. The results show the successful expression of factor FVIII gene and its product in the mouse mammary glands.
authors.affiliation: 1 Department of Microbiology, Shahr-e-Qods Branch, Islamic Azad University , Tehran, Iran .
authors.forename: Tahar
authors.initials: T
authors.lastname: Mohammadian
authors.forename: Hossein
authors.initials: H
authors.lastname: Rassi
mesh_terms: D000818:Animals; D005169:Factor VIII / Q000235:genetics* / Q000378:metabolism*; D015870:Gene Expression / Q000235:genetics; D005822:Genetic Vectors / Q000235:genetics; D006467:Hemophilia A / Q000235:genetics / Q000378:metabolism; D006801:Humans; D008321:Mammary Glands, Animal / Q000378:metabolism*; D051379:Mice; D008807:Mice, Inbred BALB C; D008822:Mice, Transgenic / Q000235:genetics*; D008892:Milk / Q000378:metabolism; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics
chemical_list: D005169:Factor VIII