Cloning and expression of the cDNA for canine tumor necrosis factor-α in E. coli

K. Zucker, P. Lu, L. Fuller, D. Asthana, V. Esquenazi, J. Miller

Research output: Contribution to journalArticle

25 Citations (Scopus)

Abstract

We have utilized the reverse transcription polymerase chain reaction (RT- PCR) to clone the protein coding region of canine tumor necrosis factor (TNF- α) cDNA. The gene displays 90% sequence homology to the corresponding human TNF-α cDNA. The predicted initial translation product is 233 amino acids and shows 88% homology to the human counterpart, and 92% homology with the human putative mature TNF-α protein. The canine TNF-α clone was used to engineer bacteria to express large amounts of the mature form of recombinant protein. A monoclonal antibody against human TNF-α cross-reacted with canine rTNF-α using Western blot and ELISA analysis. The purified canine rTNF-α had a cytotoxic effect on WEHI 164 clone 13 cells as well as increasing the cell surface expression of major histocompatibility class II antigens on canine kidney cortical cell line (MDCK) in vitro. The availability of canine rTNF- α will allow further studies on its role in immunoregulatory mechanisms in the canine transplantation model, both by itself and in conjunction with the already available canine specific recombinant interferon-γ.

Original languageEnglish
Pages (from-to)191-196
Number of pages6
JournalLymphokine and Cytokine Research
Volume13
Issue number3
StatePublished - Jan 1 1994
Externally publishedYes

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Canidae
Organism Cloning
Complementary DNA
Tumor Necrosis Factor-alpha
Escherichia coli
Clone Cells
Histocompatibility Antigens Class II
Sequence Homology
Recombinant Proteins
Interferons
Open Reading Frames
Reverse Transcription
Transplantation
Western Blotting
Enzyme-Linked Immunosorbent Assay
Monoclonal Antibodies
Bacteria
Kidney
Amino Acids
Cell Line

ASJC Scopus subject areas

  • Immunology
  • Hematology

Cite this

Zucker, K., Lu, P., Fuller, L., Asthana, D., Esquenazi, V., & Miller, J. (1994). Cloning and expression of the cDNA for canine tumor necrosis factor-α in E. coli. Lymphokine and Cytokine Research, 13(3), 191-196.

Cloning and expression of the cDNA for canine tumor necrosis factor-α in E. coli. / Zucker, K.; Lu, P.; Fuller, L.; Asthana, D.; Esquenazi, V.; Miller, J.

In: Lymphokine and Cytokine Research, Vol. 13, No. 3, 01.01.1994, p. 191-196.

Research output: Contribution to journalArticle

Zucker, K, Lu, P, Fuller, L, Asthana, D, Esquenazi, V & Miller, J 1994, 'Cloning and expression of the cDNA for canine tumor necrosis factor-α in E. coli', Lymphokine and Cytokine Research, vol. 13, no. 3, pp. 191-196.
Zucker K, Lu P, Fuller L, Asthana D, Esquenazi V, Miller J. Cloning and expression of the cDNA for canine tumor necrosis factor-α in E. coli. Lymphokine and Cytokine Research. 1994 Jan 1;13(3):191-196.
Zucker, K. ; Lu, P. ; Fuller, L. ; Asthana, D. ; Esquenazi, V. ; Miller, J. / Cloning and expression of the cDNA for canine tumor necrosis factor-α in E. coli. In: Lymphokine and Cytokine Research. 1994 ; Vol. 13, No. 3. pp. 191-196.
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