Folia Microbiologica

, Volume 45, Issue 2, pp 114–120 | Cite as

Construction of promoter-probe shuttle vectors forEscherichia coli and corynebacteria on the basis of promoterless α-amylase gene

  • J. Ugorčáková
  • G. Bukovská
  • J. Timko


We constructed new promoter-probe vectors forE. coli and corynebacteria based on the promoterless α-amylase gene originating fromBacillus subtilis. Vectors pJUPAE1 and pJUPAE2 are suitable for isolation of transcriptionally active fragments from plasmids, phages or genomic DNA α-Amylase activity can be easily visually detected on agar plates containing a chromogenic substrate, or by direct measurement of α-amylase activity.


Chromogenic Substrate Amino Acid Biosynthetic Pathway Brevibacterium Lactofermentum Ptac Promoter Industrial Production Strain 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.


Unable to display preview. Download preview PDF.

Unable to display preview. Download preview PDF.


  1. Ausubel F.M., Brent R., Kingston R.E., Moore D.O., Seidmann J.S., Smith J., Struhl K.:Current Protocols in Molecular Biology. John Wiley, New York 1987.Google Scholar
  2. Barák I., Koptides M., Jucovič M., Šišová M., Timko J.: Construction of promoter-probe shuttle vector forE. coli and brevibacteria.Gene95, 133–135 (1990).PubMedCrossRefGoogle Scholar
  3. Bathe B., Kalinowski J., Pühler A.: A physical and genetic map of theCorynebacterium glutamicum ATCC 13032 chromosome.Mol. Gen. Genet.252, 255–265 (1996).PubMedGoogle Scholar
  4. Biely P., Mislovičová D., Markovič O., Kaláč V.: A new chromogenic substrate for assay and detection of α-amylase.Anal. Biochem.172, 176–179 (1988).PubMedCrossRefGoogle Scholar
  5. Bukovská B., Slugeňová M., Timko J.: The α-amylase gene from an industrial production strain ofBacillus subtilis, pp. 197–209 in J. Balan (Ed.):Proc. Symp. Metabolism and Enzymology of Nucleic Acids Including Gene and Protein Engineering 7, Bratislava 1991.Google Scholar
  6. Cadenas R.F., Martín J.F., Gil J.A.: Construction and characterization promoter-probe vectors for corynebacteria using the kanamycin-resistance reporter gene.Gene98, 117–121 (1991).PubMedCrossRefGoogle Scholar
  7. Cadenas R.F., Fernandez-Gonzalez C., Martín J.F., Gil J.A.: Construction of new cloning vectors forBrevibacterium lactofermentum.FEMS Microbiol. Lett.137, 63–68 (1996).PubMedCrossRefGoogle Scholar
  8. Correia A., Martín J.F., Castro J.M.: Pulsed-field gel electrophoresis analysis of the genome of amino acid-producing corynebacteria: chromosome sizes and diversity of restriction patterns.Microbiology (UK)140, 2841–2847 (1994).CrossRefGoogle Scholar
  9. Guerrero C., Mateos L.M., Malumbres M., Martín J.F.: Directed mutagenesis of regulatory palindromic sequence upstream from theBrevibacterium lactofermentum tryptophan operon.Gene138, 35–41 (1994).PubMedCrossRefGoogle Scholar
  10. Haynes J.A., Britz M.L.: The effect of gowth conditions ofCorynebacterium glutamicum on the transformation frequency obtained by electroporation.J. Gen. Microbiol.136, 255–263 (1990).Google Scholar
  11. Jucovič M., Ugorčáková J., Barák I., Timko J.: Transformation of amino acid-producing brevibacteria with recombinant plasmids carrying the α-amylase gene.Biológia4, 281–288 (1991).Google Scholar
  12. Koptides M., Barák I., Šišová M., Baloghová E., Ugorčáková J., Timko J.: Characterization of bacteriophage BFK20 fromBrevibacterium flavum.J. Gen. Microbiol.138, 1387–1391 (1992).PubMedGoogle Scholar
  13. Koptides M., Ugorčáková J., Baloghová E., Bukovská G., Timko J.: Characterization and sequence analysis of the F2 promoter from corynephage BFK20.Acta Virol.38, 223–228 (1994).PubMedGoogle Scholar
  14. Malumbres M., Gil J.A., Martín J.F.: Codon preference in corynebacteria.Gene134, 15–24 (1993).PubMedCrossRefGoogle Scholar
  15. Martín J.F., Cadenas R.F., Malumbres M., Mateos L.M., Guerrero C., Gil J.A.: Construction and utilization of promoter-probe and expression vectors in corynebacteria. Characterization of corynebacterial promoters, pp. 283–292 in H. Heslot, J. Davies, J. Florent, L. Bobichon, G. Durant, L. Penasse (Eds):Proc. 6th Internat. Symp. Genetics of Industrial Microorganisms. Société Francaise de Microbiologie, Strasbourg 1990.Google Scholar
  16. Morinaga Y., Tsuchiya M., Miwa K., Sano K.: Expression ofEscherichia coli promoters inBrevibacterium lactofermentum using the shuttle vector pEB003.J. Biotechnol.5, 305–312 (1987).CrossRefGoogle Scholar
  17. Oguiza J.A., Marcos A.T., Malumbres M., Martín J.F.: Multiple σ factors inBrevibacterium lactofermentum: characterization ofsigA andsigB.J. Bacteriol.178, 550–553 (1996).PubMedGoogle Scholar
  18. Pátek M., Eikmans B.J., Pátek J., Sahm H.: Promoters fromCorynebacterium glutamicum: cloning, molecular analysis and search for consensus motif.Microbiology142, 1297–1309 (1996).PubMedCrossRefGoogle Scholar
  19. Rašlík I.: Cloning and characterization of fragments with promoter activity.BSc Thesis. Faculty of Science, Cómenius University, Bratislava 1996.Google Scholar
  20. Sambrook J., Fritsch E.F., Maniatis T.:Molecular Cloning. A Laboratory Manual, 2nd ed. Cold Spring Harbor Laboratory Press, Cold Spring Harbor 1989.Google Scholar
  21. Santamaria R.I., Gil J.A., Mesas J.M., Martín J.F.: Characterization of an endogenous plasmid and development of cloning vectors and a transformation system inBrevibacterium lactofermentum.J. Gen. Microbiol.130, 2237–2246 (1984).Google Scholar
  22. Slugeňová M., Bukovská G., Timko J., Zelinková E., Zelinka J.: Molecular cloning of the α-amylase gene from an industrial production strain.Biológia43, 1055–1067 (1988).Google Scholar
  23. Ugorčáková J., Jucovič M., Bukovská G., Timko J.: Construction and characterization of new corynebacterial plasmids carrying the α-amylase gene.Folia Microbiol.41, 10–14 (1996).CrossRefGoogle Scholar
  24. Vertés A.A., Harakeyama K., Inui M., Kobayashi M., Kurusu Y., Yukawa H.: Replacement recombination in coryneform bacteria: high efficiency integration requirement for non-methylated plasmid DNA.Biosci. Biotech. Biochem.57, 2036–2038 (1993).Google Scholar

Copyright information

© Institute of Microbiology, Academy of Sciences of the Czech Republic 2000

Authors and Affiliations

  • J. Ugorčáková
    • 1
  • G. Bukovská
    • 1
  • J. Timko
    • 1
  1. 1.Institute of Molecular BiologySlovak Academy of SciencesBratislavaSlovakia

Personalised recommendations