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. 1988 Nov;54(11):2672–2676. doi: 10.1128/aem.54.11.2672-2676.1988

Cloning and expression of a Bacteroides succinogenes mixed-linkage beta-glucanase (1,3-1,4-beta-D-glucan 4-glucanohydrolase) gene in Escherichia coli.

J E Irvin 1, R M Teather 1
PMCID: PMC204354  PMID: 3145707

Abstract

A pseudorandom genomic library of Bacteroides succinogenes DNA, cloned into pUC8 in Escherichia coli, was screened for beta-glucanase activity on 0.1% lichenan plates. Six high-activity clones, containing identical 5.2-kilobase inserts of B. succinogenes DNA, were obtained. The clones exhibited activity solely on beta-glucan substrates containing beta-(1----3)(1----4) linkages, thus manifesting a specific fibrolytic enzyme previously unrecognized in B. succinogenes. A subclone (pJI10) of the original insert (1.35 kilobases in size) expressed full beta-glucanase activity under control of its own promoter. The expression of beta-glucanase in pJI10 appeared subject to catabolite regulation by glucose. Detailed analysis of enzyme activity in the parental and deleted derivatives, subcloned into pUC18 and pUC19, suggested that the apparent glucose repression was an artifact arising as a consequence of interactions with the lac transcriptional unit in the plasmid vector.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Birnboim H. C., Doly J. A rapid alkaline extraction procedure for screening recombinant plasmid DNA. Nucleic Acids Res. 1979 Nov 24;7(6):1513–1523. doi: 10.1093/nar/7.6.1513. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Cantwell B. A., McConnell D. J. Molecular cloning and expression of a Bacillus subtilis beta-glucanase gene in Escherichia coli. Gene. 1983 Aug;23(2):211–219. doi: 10.1016/0378-1119(83)90053-7. [DOI] [PubMed] [Google Scholar]
  3. Clewell D. B. Nature of Col E 1 plasmid replication in Escherichia coli in the presence of the chloramphenicol. J Bacteriol. 1972 May;110(2):667–676. doi: 10.1128/jb.110.2.667-676.1972. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Feinberg A. P., Vogelstein B. A technique for radiolabeling DNA restriction endonuclease fragments to high specific activity. Anal Biochem. 1983 Jul 1;132(1):6–13. doi: 10.1016/0003-2697(83)90418-9. [DOI] [PubMed] [Google Scholar]
  5. Forsberg C. W., Crosby B., Thomas D. Y. Potential for manipulation of the rumen fermentation through the use of recombinant DNA techniques. J Anim Sci. 1986 Jul;63(1):310–325. doi: 10.2527/jas1986.631310x. [DOI] [PubMed] [Google Scholar]
  6. Gough G. W., Lilley D. M. DNA bending induced by cruciform formation. Nature. 1985 Jan 10;313(5998):154–156. doi: 10.1038/313154a0. [DOI] [PubMed] [Google Scholar]
  7. Hinchliffe E. Cloning and expression of a Bacillus subtilis Endo-1,3-1,4-beta-D-glucanase gene in Escherichia coli K12. J Gen Microbiol. 1984 May;130(5):1285–1291. doi: 10.1099/00221287-130-5-1285. [DOI] [PubMed] [Google Scholar]
  8. Sipat A., Taylor K. A., Lo R. Y., Forsberg C. W., Krell P. J. Molecular cloning of a xylanase gene from Bacteroides succinogenes and its expression in Escherichia coli. Appl Environ Microbiol. 1987 Mar;53(3):477–481. doi: 10.1128/aem.53.3.477-481.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Smith C. J., Hespell R. B. Prospects for development and use of recombinant deoxyribonucleic acid techniques with ruminal bacteria. J Dairy Sci. 1983 Jul;66(7):1536–1546. doi: 10.3168/jds.S0022-0302(83)81970-5. [DOI] [PubMed] [Google Scholar]
  10. Taylor K. A., Crosby B., McGavin M., Forsberg C. W., Thomas D. Y. Characteristics of the endoglucanase encoded by a cel gene from Bacteroides succinogenes expressed in Escherichia coli. Appl Environ Microbiol. 1987 Jan;53(1):41–46. doi: 10.1128/aem.53.1.41-46.1987. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Teather R. M., Müller-Hill B., Abrutsch U., Aichele G., Overath P. Amplification of the lactose carrier protein in Escherichia coli using a plasmid vector. Mol Gen Genet. 1978 Feb 27;159(3):239–248. doi: 10.1007/BF00268260. [DOI] [PubMed] [Google Scholar]
  12. Teather R. M., Wood P. J. Use of Congo red-polysaccharide interactions in enumeration and characterization of cellulolytic bacteria from the bovine rumen. Appl Environ Microbiol. 1982 Apr;43(4):777–780. doi: 10.1128/aem.43.4.777-780.1982. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. Vieira J., Messing J. The pUC plasmids, an M13mp7-derived system for insertion mutagenesis and sequencing with synthetic universal primers. Gene. 1982 Oct;19(3):259–268. doi: 10.1016/0378-1119(82)90015-4. [DOI] [PubMed] [Google Scholar]
  14. Wu H. M., Crothers D. M. The locus of sequence-directed and protein-induced DNA bending. Nature. 1984 Apr 5;308(5959):509–513. doi: 10.1038/308509a0. [DOI] [PubMed] [Google Scholar]
  15. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]

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