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Current IssueEditorial BoardOnline First ArticlesArchive

volume 47 issue 1 (february 2013) : 61-65

DIFFERENTIAL EXPRESSION PROFILING OF RECOMBINANT BOVINE INHIBIN-ALPHA AT REDUCED TEMPERATURE

A
Anuradha Bhardwaj*
V
Varij Nayan1
S
S. De2
S
S.L. Goswami
1Genomics and Molecular Biology Lab., Animal Biotechnology Centre, National Dairy Research Institute, Karnal- 132 001, India

  • Submitted|

  • First Online |

  • doi

Cite article:- Bhardwaj* Anuradha, Nayan1 Varij, De2 S., Goswami S.L. (2025). DIFFERENTIAL EXPRESSION PROFILING OF RECOMBINANT BOVINE INHIBIN-ALPHA AT REDUCED TEMPERATURE. Indian Journal of Animal Research. 47(1): 61-65. doi: .

ABSTRACT

The molecular biology approach for production of recombinant applied products in heterologous systems such as prokaryotes and eukaryotes is becoming a method of choice in the post genomics era. Many heterologous expression systems are available such as insects, yeast, mammalian cells however, Escherichia coli is most desirable host but the main drawback associated with expression of recombinant proteins in E. coli is production of insoluble and/or nonfunctional proteins. Earlier, effort was made to produce recombinant inhibin alpha protein to be used as fecundity vaccine in small ruminants. The bovine inhibin alpha immunogenic region of about 134 amino acid region was targeted for high level expression in E.coli but expression at 370C resulted in high level of inclusion bodies though yield of purified protein was satisfactory. In the present study, reduced temperature (22°C) with different time intervals were investigated for the expression of soluble recombinant bINH-a aiming at improving and facilitating recombinant bINH-a protein production and purification from E. coli and to characterize it by immunological techniques.

REFERENCES

  1. Bhardwaj A., Nayan V., Parvati, Mamta and Gupta A.K. (2012a) Inhibin: A role for fecundity augmentation in farm animals. Asian J. Anim. Vete. Adva. 7 : 771-789. 
  2. Bhardwaj A., Nayan V., Yadav P., De S., Datta T.K. and Goswami S.L. (2012b) Heterologous Expression and Characterization of Indian Sahiwal Cattle (Bos indicus) Alpha Inhibin, Anim. Biotechnol. 23 : 71-88.
  3. Cabilly, S. (1989) Growth at sub-optimal temperatures allows the production of functional, antigen-binding Fab fragments in Escherichia coli. Gene 85: 553–557.
  4. de Jong F.H. and Sharpe R.M. (1976) Evidence for inhibin-like activity in bovine follicular fluid. Nature 263:71-72.
  5. Escher, A., O’Kane, D. J., Lee, J., and Szalay, A. A. (1989) Bacterial luciferase ab fusion protein is fully active as a monomer and highly sensitive in vivo to elevated temperatures. Proc. Natl. Acad. Sci. USA 86: 6528–6532.
  6. Forage R.G., Brown R.W., Oliver K.J., Atrache B.T., Devine P.L., Hudson G.C., Goss N.H., Bertram K.C., Tolstoshev P., Robertson D.M., de Kretser D.M., Doughton B., Burger H.G., Findlay J.K. (1987) Immunization against an inhibin subunit produced by recombinant DNA techniques results in increased ovulation rate in sheep. J. Endocrinol. 114: Rl-R4.
  7. Jones, P.G., VanBogelen, R.A., and Neidhardt, F.C. (1987) Induction of proteins in response to low temperatures in Escherichia coli. J. Bacteriol. 169: 2092–2095.
  8. Kim K.D., Knight P.G. and Savva D. (1991) Expression of bovine inhibin beta subunit in E.coli. Int. J. Biochem. 23(11):1307-1313.
  9. Medan, M.S., Arai, K.V., Watanabe, G. and Taya, K. (2007) Inhibin: Regulation of reproductive function and. practical use in females. Anim. Sci. J., 78: 16-27.
  10. Pangas S.A. and Woodruff T.K. (2002) Production and purification of recombinant human inhibin and activin. J. Endocrinol. 172:199–210.
  11. Sambrook J. and Russell D.W. (2001) Molecular Cloning: A Laboratory Manual. 3rd ed. Cold Spring Harbor Laboratory Press.
  12. Schein, C.H. and Noteborn, M.H. M. (1988) Formation of soluble recombinant proteins in Escherichia coli is favored by lower growth temperatures. Bio/Technology 6: 291–294.
  13. Schumann, W. and Ferreira, L.C.S. (2004) Production of recombinant proteins in E.coli, Gene. Mol. Biol. 27 : 442- 453. 
  14. Sørensen, H.P. and Mortensen, K.K. (2005) Soluble expression of recombinant proteins in the cytoplasm of Escherichia coli. Microb. Cell Fact., 4(1):1.
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Indian Journal of Animal Research
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    volume 47 issue 1 (february 2013) : 61-65

    DIFFERENTIAL EXPRESSION PROFILING OF RECOMBINANT BOVINE INHIBIN-ALPHA AT REDUCED TEMPERATURE

    A
    Anuradha Bhardwaj*
    V
    Varij Nayan1
    S
    S. De2
    S
    S.L. Goswami
    1Genomics and Molecular Biology Lab., Animal Biotechnology Centre, National Dairy Research Institute, Karnal- 132 001, India

    • Submitted|

    • First Online |

    • doi

    Cite article:- Bhardwaj* Anuradha, Nayan1 Varij, De2 S., Goswami S.L. (2025). DIFFERENTIAL EXPRESSION PROFILING OF RECOMBINANT BOVINE INHIBIN-ALPHA AT REDUCED TEMPERATURE. Indian Journal of Animal Research. 47(1): 61-65. doi: .

    ABSTRACT

    The molecular biology approach for production of recombinant applied products in heterologous systems such as prokaryotes and eukaryotes is becoming a method of choice in the post genomics era. Many heterologous expression systems are available such as insects, yeast, mammalian cells however, Escherichia coli is most desirable host but the main drawback associated with expression of recombinant proteins in E. coli is production of insoluble and/or nonfunctional proteins. Earlier, effort was made to produce recombinant inhibin alpha protein to be used as fecundity vaccine in small ruminants. The bovine inhibin alpha immunogenic region of about 134 amino acid region was targeted for high level expression in E.coli but expression at 370C resulted in high level of inclusion bodies though yield of purified protein was satisfactory. In the present study, reduced temperature (22°C) with different time intervals were investigated for the expression of soluble recombinant bINH-a aiming at improving and facilitating recombinant bINH-a protein production and purification from E. coli and to characterize it by immunological techniques.

    REFERENCES

    1. Bhardwaj A., Nayan V., Parvati, Mamta and Gupta A.K. (2012a) Inhibin: A role for fecundity augmentation in farm animals. Asian J. Anim. Vete. Adva. 7 : 771-789. 
    2. Bhardwaj A., Nayan V., Yadav P., De S., Datta T.K. and Goswami S.L. (2012b) Heterologous Expression and Characterization of Indian Sahiwal Cattle (Bos indicus) Alpha Inhibin, Anim. Biotechnol. 23 : 71-88.
    3. Cabilly, S. (1989) Growth at sub-optimal temperatures allows the production of functional, antigen-binding Fab fragments in Escherichia coli. Gene 85: 553–557.
    4. de Jong F.H. and Sharpe R.M. (1976) Evidence for inhibin-like activity in bovine follicular fluid. Nature 263:71-72.
    5. Escher, A., O’Kane, D. J., Lee, J., and Szalay, A. A. (1989) Bacterial luciferase ab fusion protein is fully active as a monomer and highly sensitive in vivo to elevated temperatures. Proc. Natl. Acad. Sci. USA 86: 6528–6532.
    6. Forage R.G., Brown R.W., Oliver K.J., Atrache B.T., Devine P.L., Hudson G.C., Goss N.H., Bertram K.C., Tolstoshev P., Robertson D.M., de Kretser D.M., Doughton B., Burger H.G., Findlay J.K. (1987) Immunization against an inhibin subunit produced by recombinant DNA techniques results in increased ovulation rate in sheep. J. Endocrinol. 114: Rl-R4.
    7. Jones, P.G., VanBogelen, R.A., and Neidhardt, F.C. (1987) Induction of proteins in response to low temperatures in Escherichia coli. J. Bacteriol. 169: 2092–2095.
    8. Kim K.D., Knight P.G. and Savva D. (1991) Expression of bovine inhibin beta subunit in E.coli. Int. J. Biochem. 23(11):1307-1313.
    9. Medan, M.S., Arai, K.V., Watanabe, G. and Taya, K. (2007) Inhibin: Regulation of reproductive function and. practical use in females. Anim. Sci. J., 78: 16-27.
    10. Pangas S.A. and Woodruff T.K. (2002) Production and purification of recombinant human inhibin and activin. J. Endocrinol. 172:199–210.
    11. Sambrook J. and Russell D.W. (2001) Molecular Cloning: A Laboratory Manual. 3rd ed. Cold Spring Harbor Laboratory Press.
    12. Schein, C.H. and Noteborn, M.H. M. (1988) Formation of soluble recombinant proteins in Escherichia coli is favored by lower growth temperatures. Bio/Technology 6: 291–294.
    13. Schumann, W. and Ferreira, L.C.S. (2004) Production of recombinant proteins in E.coli, Gene. Mol. Biol. 27 : 442- 453. 
    14. Sørensen, H.P. and Mortensen, K.K. (2005) Soluble expression of recombinant proteins in the cytoplasm of Escherichia coli. Microb. Cell Fact., 4(1):1.
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