Isolation and culture of bovine mammary epithelial cells and establishment of gene transfection conditions in the cells.

Bovine mammary epithelial cells (BMEC) were isolated as acinous fragments from a mammary gland of a lactating cow. They grew well on plastic substratum, showed the characteristic cobblestone morphology of epithelial cells, and secreted alpha s1-, beta-, and kappa-caseins even when grown on plastic substratum. A plasmid containing the bacterial chloramphenicol acetyl transferase (CAT) gene was transfected to the isolated BMEC by calcium phosphate precipitation and electroporation methods. The transfection efficiency of BMEC by the calcium phosphate method was greatly improved by post-transfection osmotic shock with glycerol or polyethylene glycol. An about 700 bp DNA fragment containing 5'-flanking sequence of bovine alpha s1-casein gene showed promoter activity in the transfected BMEC. The primary culture of BMEC might be useful for studies on regulation of bovine milk-protein gene expression.