Effect of amplification of dhfr and lac Z genes on growth and β-galactosidase expression in suspension cultures of recombinant CHO cells

Man Bock Gu, Jeffrey A. Kern, Paul Todd, Dhinakar S. Kompala

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Abstract

Studies were conducted to characterize the effect of gene amplification and foreign gene expression on recombinant CHO cell growth. Chinese hamster ovary (CHO) cells were transfected with an expression vector containing the gene for dihydrofolate reductase (dhfr) and the gene for human β-interferon (β-IFN) or the lac Z gene which codes for β-galactosidase (β-gal). The recombinant genes in these CHO cells were amplified stepwise by growth in 0, 10-7, and 10-6 M methotrexate (MTX), and the β-gal expressing cells were adapted to suspension culture. Flow cytometric methods (FCM) were used to measure the distribution of amplified dhfr gene content and foreign β-gal gene expression in the cell populations. A biochemical assay for β-gal was also used. Beta-gal expression was found to increase with increasing gene amplification. The growth rate of recombinant CHO cells at 10-7 M MTX was found to be 20% lower than that of recombinant CHO cells in MTX-free medium, and the cell growth rate at 10-6 M MTX was 20% lower than that of recombinant CHO cells at 10-7 M MTX. There was no effect of 10-5 M MTX on the growth of CHO-DG44 (dhfr-) cells. The reduction of growth rate in recombinant CHO cells is therefore thought to be mainly due to the effect of dhfr and foreign gene amplification and increased β-galactosidase expression.

Original languageEnglish
Pages (from-to)237-245
Number of pages9
JournalCytotechnology
Volume9
Issue number1-3
DOIs
Publication statusPublished - 1992 Feb 1
Externally publishedYes

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Keywords

  • dhfr
  • gene amplification
  • growth rate reduction
  • recombinant CHO cells
  • recombinant gene expression
  • suspension culture

ASJC Scopus subject areas

  • Clinical Biochemistry
  • Cell Biology
  • Biotechnology

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