Metabolic burden in recombinant CHO cells: effect of dhfr gene amplification and lacZ expression

Man Bock Gu, Paul Todd, Dhinakar S. Kompala

Research output: Contribution to journalArticlepeer-review

36 Citations (Scopus)

Abstract

Foreign protein production levels in two recombinant Chinese hamster ovary (CHO) cell lines were compared in cells transfected with different expression vectors. One vector pNL1 contained the gene for neomycin resistance (neor) and the lacZ gene which codes for intracellular β-galactosidase, with both genes controlled by the constitutive simian virus (SV40) promoter. The other vector CDβG contained the amplifiable dhfr gene and lacZ gene, controlled by the constitutive SV40 and cytomegalovirus (CMV) promoters, respectively. Cell growth and β-galactosidase expression were compared quantitatively after cells were selected in different concentrations of the neomycin analog G418 and methotrexate, respectively. A 62% reduction in growth rate occurred in recombinant CHO cells in which the lacZ and dhfr genes were highly amplified and expressed. In contrast, the combined effects of the unamplified neor gene and lacZ gene expression on the growth kinetics were small. Any metabolic burden caused by lacZ gene expression, which was evaluated separately from the effect of neor gene expression, must be negligible, as higher expression of β-galactosidase (1.5×10-6 units/cell) occurred in unamplified cells compared to the cells in which lacZ was amplified by the dhfr-containing vector (3×10-7 units/cell). Thus, the main factor causing severe growth reduction ("metabolic burden") in cells containing the amplified dhfr gene system was not overexpression of β-galactosidase but dhfr and lacZ gene co-amplification and dhfr gene expression.

Original languageEnglish
Pages (from-to)159-166
Number of pages8
JournalCytotechnology
Volume18
Issue number3
DOIs
Publication statusPublished - 1995 Jan
Externally publishedYes

Keywords

  • CMV promoter
  • SV40 promoter
  • dhfr
  • foreign gene expression
  • gene amplification
  • growth rate reduction
  • metabolic burden
  • recombinant CHO cells

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Biomedical Engineering
  • Clinical Biochemistry
  • Cell Biology

Fingerprint Dive into the research topics of 'Metabolic burden in recombinant CHO cells: effect of dhfr gene amplification and lacZ expression'. Together they form a unique fingerprint.

Cite this