Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Research Paper
  • Published:

Molecular Cloning of Exo–Cellobiohydrolase I Derived from Trichoderma Reesei Strain L27

Abstract

The molecular cloning and characterization of the gene encoding exo–cellobiohydrolase I (CBHI) of Trichoderma reesei strain L27 is reported. Two adjacent HindIII genomic fragments of 1.16 kb and 2.3 kb were identified using differential hybridization techniques and were sub–cloned into plasmid pBR322. The identification of the gene encoding CBHI was determined by hybrid selection and confirmed by DNA sequence analysis. There are two introns in the genomic DNA that were identified by comparing the coding sequence with the published amino acid sequence1 and confirmed by sequencing of cDNA clones. Both introns were found to contain a 10 bp sequence, CAGCT–GACTG, that is homologous to a sequence necessary for splicing of introns in yeast2.

This is a preview of subscription content, access via your institution

Access options

Buy this article

Prices may be subject to local taxes which are calculated during checkout

Similar content being viewed by others

References

  1. Fagerstam, L.G. 1981. Cellulases from Trichoderma reesei QM9414: Enzymatic and structural properties. Doctoral Thesis, University of Uppsala, Sweden.

  2. Langford, C.J. and Gallwitz, D. 1983. Evidence for an intron-contained sequence required for the splicing of yeast polymerase II transcripts. Cell 33: 519–527.

    Article  CAS  Google Scholar 

  3. Shoemaker, S.P., Watt, K., Tsitovsky, G. and Cox, R.V. 1983. Characterization and properties of cellulases purified from Trichoderma reesei strain L27. BIO/TECHNOLOGY 1: 687–690.

    CAS  Google Scholar 

  4. Shoemaker, S.P. and Brown, R.D. Jr., 1978. Characterization of endo-1,4-β-D-glucanase purified from Trichoderma viride. Biochim. Biophys. Acta 523: 147–161.

    Article  CAS  Google Scholar 

  5. White, A.R. 1981. Visual characterization of the mode of action of the cellulase enzyme system of the fungus Trichoderma reesei. Ph. D. Thesis. University of North Carolina, Chapel Hill, NC.

  6. Wood, T.M. 1982. Enzymic and other factors affecting microbial generation of soluble sugars from cellulosic materials, p. 327–334. In: Proceedings of 1982 Research and Development Division Conference. TAPPI: Ashville, NC.

    Google Scholar 

  7. Whittle, D.J., Kilburn, D.G., Warren, R.A.J., Miller, R.C. Jr., 1982. Molecular cloning of a Cellulomonas fimi cellulase gene in Escherichia coli. Gene 17: 139–145.

    Article  CAS  Google Scholar 

  8. Cornet, P., Tronik, D., Millet, J. and Aubert, J. 1983. Cloning and expression in Escherichia coli of Clostridium thermocellum genes coding for amino acid synthesis and cellulose hydrolysis. FEMS Lett. 16: 137–141.

    Article  CAS  Google Scholar 

  9. Cornet, P., Millet, J., Beguin, P., Aubert, J. 1983. Characterization of two cel (cellulose degradation) genes of Clostridium thermocellum coding for endoglucanases. BIO/TECHNOLOGY 1: 589–594.

    CAS  Google Scholar 

  10. Wilson, D.B. and Collmer, A. 1983. Cloning and Expression of a Thermomonospora YX endocellulase gene in E. coli BIO/TECHNOLOGY 1: 594–601.

    Google Scholar 

  11. Shoemaker, S.P., Raymond, J.C., Bruner, R. 1981. Cellulases: Diversity amongst improved Trichoderma strains, p. 89–109. In: Trends in the Biology of Fermentations for Fuels and Chemicals. A. Hollaender et al. (eds.) Plenum Publishing Co., New York.

    Chapter  Google Scholar 

  12. Mount, S.M. 1982. A catalogue of splice junction sequences. Nucleic Acids Res. 10: 459–472.

    Article  CAS  Google Scholar 

  13. Dobson, M.J., Tuite, M.F., Roberts, N.A., Kingsman, A.J. and Kingsman, S.M. 1982. Conservation of high efficiency promoter sequences in Saccharomyces cerevisiae. Nucleic Acids Res. 10: 2625–2637.

    Article  CAS  Google Scholar 

  14. Chen, H.R., Dayhoff, M.O., Barker, W.C., Hunt, L.T., Yeh, L.-S., George, D.G., Orcutt, B.C. 1982. Nucleic acid sequence database IV. DNA 1: 365–374.

    Article  CAS  Google Scholar 

  15. Holland, J.P. and Holland, M.J. 1979. The primary structure of a glyceraldehyde-3-phosphate dehydrogenase gene from Saccharomyces cerevisiae. J. Biol. Chem. 254: 9839–9845.

    CAS  PubMed  Google Scholar 

  16. Holland, M.J., Holland, J.P., Thill, G.P., Jackson, A.J. 1981. The primary structures of two yeast enolase genes. J. Biol. Chem. 256: 1385–1395.

    CAS  PubMed  Google Scholar 

  17. Chirgwin, J.M., Przybyla, A.E., MacDonald, R.J. and Rutter, W.J. 1979. Isolation of biologically active ribonucleic acid from sources enriched in ribonuclease. Biochem. J. 18: 5294–5299.

    Article  CAS  Google Scholar 

  18. O'Farrell, P.H. 1975. High resolution two-dimensional electrophoresis of proteins. J. Biol. Chem. 250: 4007–4021.

    CAS  PubMed  PubMed Central  Google Scholar 

  19. Bailey, J.M. and Davidson, N. 1976. Methylmercury as a reversible denaturing agent for agarose gel electrophoresis. Anal. Biochem. 70: 75–85.

    Article  CAS  Google Scholar 

  20. Seghal, P.B. and Sagar, A.D. 1980. Heterogeneity of poly(I) · poly (C)-induced fibroblast interferon mRNA species. Nature 288: 95–97.

    Article  Google Scholar 

  21. Loenen, W.A. and Brammar, W.J. 1980. A bacteriophage lambda vector for cloning large DNA fragments made with several restriction enzymes. Gene 10: 249–259.

    Article  CAS  Google Scholar 

  22. Koths, K. and Dressler, D. 1980. The rolling circle-capsid complex as an intermediate in OX DNA replication and viral assembly. J. Biol. Chem. 255: 4328–4338.

    CAS  PubMed  Google Scholar 

  23. Hohn, B. 1979. In vitro packaging of λ and cosmid DNA, p. 299–309. In: Methods in Enzymology, vol. 68. R. Wu (ed.) Academic Press, New York.

    Google Scholar 

  24. Maniatis, T., Fritsch, E.F. and Sambrook, J. 1982. Molecular cloning: A laboratory manual. Cold Spring Harbor Laboratory, Cold Spring Harbor, New York.

    Google Scholar 

  25. Meselson, M. and Yuan, R. 1968. DNA restriction enzyme from E. coli. Nature 217: 1110–1114.

    Article  CAS  Google Scholar 

  26. Clarke, L. and Carbon, J. 1976. A colony bank containing synthetic ColEl hybrid plasmids representative of the entire E. coli genome. Cell 9: 91–102.

    Article  CAS  Google Scholar 

  27. Benton, W.D. and Davis, R.W. 1977. Screening λgt recombinant clones by hybridization to single plaques in situ. Science 196: 180–182.

    Article  CAS  Google Scholar 

  28. Harpold, M.M., Dobner, P.R., Evans, R.M., Bancroft, F.C. 1978. Construction and identification by positive hybridization-translation of a bacterial plasmid containing a rat growth hormone structural gene sequence. Nucleic Acids Res. 5: 2039–2053.

    Article  CAS  Google Scholar 

  29. Sanger, F., Nicklen, S. and Coulsen, A.R. 1977. DNA sequencing with chain terminating inhibitors. Proc. Natl. Acad. Sci. USA 74: 5463–5467.

    Article  CAS  Google Scholar 

  30. Messing, J., Crea, R. and Seeburg, P.H. 1981. A system for shotgun DNA sequencing. Nucleic Acids Res. 9: 309–321.

    Article  CAS  Google Scholar 

  31. Maxam, A.M. and Gilbert, W. 1977. A new method for sequencing DNA. Proc. Natl. Acad. Sci. USA 74: 560–564.

    Article  CAS  Google Scholar 

Download references

Author information

Authors and Affiliations

Authors

Rights and permissions

Reprints and permissions

About this article

Cite this article

Shoemaker, S., Schweickart, V., Ladner, M. et al. Molecular Cloning of Exo–Cellobiohydrolase I Derived from Trichoderma Reesei Strain L27. Nat Biotechnol 1, 691–696 (1983). https://doi.org/10.1038/nbt1083-691

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nbt1083-691

This article is cited by

Search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing