Levels of expression of two reporter genes cloned into SV40 or Epstein
-Barr virus (EBV) ori-containing plasmids were measured following tran
sient transfection of cell lines constitutively expressing T-antigen o
r EBV nuclear antigen 1 (EBNA1). The TSA201 and COS7 cell lines stably
produce T-antigen and support replication of the SV40 ori-containing
constructs while the 293EBNA cell line produces EBNA1 and supports rep
lication of EBV osi-containing plasmids. We found that 293EBNA cells e
xpress >25-fold more beta-galactosidase (beta Gal) per mg protein than
COS7 cells and 11-fold more beta Gal than TSA201 cells. We also demon
strate that 293EBNA cells are able to express 70-100-fold more angiote
nsin II type-1 receptor (AT1) per mg protein than COS7 or TSA201 cells
. We examined the suitability of each cell line for use in expression
cloning using a NaOH 'scrape' method as an improvement over emulsion a
utoradiography for detection, Measurable AT1 signals can be detected w
hen reporter plasmids are diluted up to 1000-fold for COS7 and TSA201
cells, and up to 80 000-fold for 293EBNA cells. These data demonstrate
that 293EBNA cells offer a significant improvement in expression clon
ing technology as compared to the conventionally used T-antigen-based
cell lines.