Serum-free transfection and selection in Chinese hamster ovary (CHO) cells.
In this report a method for transfection and selection of mammalian cells in serum-free medium is described. Chinese hamster ovary (CHO) cells were grown in serum-free medium in plastic dishes coated with one of the following attachment factors: poly-D-lysine, Cell-Tak (polyphenolic proteins extracted from the marine mussel Mytilus edulis), fibronectin or laminin. Cells grown to 80% confluence were transfected with an expression vector encoding the hygromycin resistance gene as a selectable marker and beta-galactosidase as the reporter gene. Transfectants were selected using hygromycin at a concentration of 500 micrograms/ml. Both fibronectin and laminin supported colony formation following selection in serum-free medium. However, poly-D-lysine and Cell-Tak did not. This method can, thus, be used to select for clones producing a recombinant product in cells that are growing in serum-free medium from the onset to provide a better system from which to purify proteins.
Zaworski
P G
PG
Upjohn Company, Kalamazoo, MI 49007.
Gill
G S
GS
D000818:Animals; D001769:Blood; D016466:CHO Cells / Q000378:metabolism*; D002448:Cell Adhesion; D002934:Cinnamates; D006224:Cricetinae; D003470:Culture Media; D004351:Drug Resistance / Q000235:genetics; D005353:Fibronectins; D006921:Hygromycin B / Q000031:analogs & derivatives / Q000494:pharmacology; D007797:Laminin; D008565:Membrane Proteins; D011107:Polylysine; D011994:Recombinant Proteins / Q000096:biosynthesis*; D014162:Transfection; D001616:beta-Galactosidase / Q000235:genetics
C056580:Cell-Tak; D002934:Cinnamates; D003470:Culture Media; D005353:Fibronectins; D007797:Laminin; D008565:Membrane Proteins; D011994:Recombinant Proteins; D011107:Polylysine; D006921:Hygromycin B; C026273:hygromycin A; D001616:beta-Galactosidase