Development of a high-efficiency method for gene marking of Dunning prostate cancer cell lines with the enzyme beta-galactosidase.

Journal Article (Journal Article)

Although the bacterial enzyme beta-galactosidase has been used as a reporter gene in a variety of mammalian systems; the variability and instability of its expression has limited its use. Transfection of Dunning rat prostatic cell lines with beta-galactosidase expression plasmids resulted in 5-10% of cells expressing the enzyme transiently, and < 5% of G418-resistant clones showing any level of expression. To address this problem, we developed a labeling protocol using a replication defective retrovirus containing a beta-galactosidase expression cassette. Between 30-50% of cells transduced expressed high levels of this enzyme. Homogeneous cell populations were isolated by subsequent fluorescence-activated cell sorting, using a fluorescent beta-galactosidase substrate. Using a modification of standard staining procedures, small metastatic foci of cells expressing beta-galactosidase in mouse lung tissue were detected with high sensitivity. This method has several advantages over standard transfection protocols, including the expedient and efficient transfer of the beta-galactosidase gene and the stability of its expression in a variety of Dunning sublines.

Full Text

Duke Authors

Cited Authors

  • Rinker-Schaeffer, CW; Wharam, JF; Simons, J; Isaacs, JT

Published Date

  • July 1996

Published In

Volume / Issue

  • 29 / 1

Start / End Page

  • 60 - 64

PubMed ID

  • 8685057

International Standard Serial Number (ISSN)

  • 0270-4137

Digital Object Identifier (DOI)

  • 10.1002/(SICI)1097-0045(199607)29:1<60::AID-PROS9>3.0.CO;2-M

Language

  • eng

Conference Location

  • United States