Expression of immunoglobulin genes tandem in eukaryotic cells under the control of T7 bacteriophage RNA polymerase

Sergey M. Deyev, Andre Lieber, Boris V. Radko, Oleg L. Polanovsky

Research output: Contribution to journalArticle

5 Citations (Scopus)

Abstract

A tandem of recombinant mouse/human immunoglobulin (Ig) genes was constructed and inserted into the plasmid pGEM1 under the control of T7 phage RNA polymerase promoter. Sp2/0 lymphoid cell line and Chinese Hamster Ovary (CHO) cells were used as the targets for gene transfection. Both cell lines contained in their genomes a T7 RNA polymerase gene modified with a nuclear-located signal derived from SV40 large T-antigen. Cell lines transfected with the gene tandem effectively synthesized mRNA (up to 9 × 103 bp) that hybridized with ε-and κ{script}-gene probes. Separate transcripts corresponding to mRNAs of individual heavy and light chains were not detected in either transfected cell line. It follows from these data that transcription in the transfected cells is controlled mainly by the T7 phage polymerase promoter. Both lymphoid and nonlymphoid cell lines transfected with the gene tandem synthesized the ε-heavy (70 kDa) and κ{script}-light (25 kDa) Ig polypeptide chains. Production of chimeric antibodies by the myeloma Sp2/0 cells was higher than that by the CHO cells. Individual clones synthesized up to 150 ng/mL chimeric IgE. However, only lymphoid Sp2/0 cells were capable of efficient secretion of the recombinant antibodies. The mechanism of translation of mRNA synthesized in eukaryotic cells by T7 phage RNA polymerase is discussed.

Original languageEnglish
Pages (from-to)143-155
Number of pages13
JournalApplied Biochemistry and Biotechnology
Volume47
Issue number2-3
DOIs
Publication statusPublished - May 1994
Externally publishedYes

Fingerprint

Immunoglobulin Genes
Bacteriophages
Eukaryotic Cells
RNA
Immunoglobulins
Bacteriophage T7
Genes
Cell Line
Cells
Cricetulus
Ovary
Immunoglobulin Subunits
DNA-Directed RNA Polymerases
Lymphocytes
Antibodies
Polyomavirus Transforming Antigens
Messenger RNA
Light
Viral Tumor Antigens
Protein Biosynthesis

Keywords

  • CHO cell
  • gene tandem expression
  • phage T7 RNA polymerase
  • Recombinant antibodies
  • Sp2/0 cell

ASJC Scopus subject areas

  • Biotechnology
  • Bioengineering
  • Biochemistry
  • Applied Microbiology and Biotechnology
  • Molecular Biology

Cite this

Expression of immunoglobulin genes tandem in eukaryotic cells under the control of T7 bacteriophage RNA polymerase. / Deyev, Sergey M.; Lieber, Andre; Radko, Boris V.; Polanovsky, Oleg L.

In: Applied Biochemistry and Biotechnology, Vol. 47, No. 2-3, 05.1994, p. 143-155.

Research output: Contribution to journalArticle

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