Expression of human serum albumin in milk of transgenic mice using goat β-casein/human serum albumin fusion gene

Abstract

The gene encoding human serum albumin (HSA) was cloned from human liver cDNA library by PCR. The HSA cDNA in size of 2,176 bp, including 1,830 bp of open reading frame, was cloned into the plasmid carried with the 5′ flanking sequence of goat β-casein gene (-4,044 to +2,025 bp) to get a tissue specific expression vector in mammary gland named pGB562/HSA (12.5 kb). A 9.6 kb DNA fragment in which the sequence is in order of goat β-casein gene regulatory sequence, HSA cDNA and SV40 polyadenylation signals was isolated from the pGB562/HSA by SacI and DraIII cutting, and used to microinject into the pronuclei of mouse fertilized eggs to produce transgenic mice. Three transgenic mice (2 female and 1 male) were identified by PCR and dot Southern blot analysis. The copy numbers of integrated transgene were more than 10 copies in line #21 and #26 as well as over 50 copies in line #31 of transgenic mice. HSA protein collected from the milk of lactating transgenic mice was confirmed by immuno-detection of Western and slot blot. The concentrations of HSA in the milk were from 0.05 to 0.4 mg/ml. An obvious antigen and antibody conjugate could be observed in immunohistochemical stain of mammary gland tissue from lactating day 11 of HSA transgenic mice. The transmission of transgene and its expression was recognized according to the results of RT-PCR and sequences analyses of their progeny.