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http://dx.doi.org/10.5423/PPJ.2009.25.2.136

Generation of a Constitutive Green Fluorescent Protein Expression Construct to Mark Biocontrol Bacteria Using P43 Promoter from Bacillus subtilis  

Kong, Hyun-Gi (Department of Applied Biology, Dong-A University)
Choi, Ki-Hyuck (Department of Applied Biology, Dong-A University)
Heo, Kwang-Ryool (Department of Applied Biology, Dong-A University)
Lee, Kwang-Youll (Department of Applied Biology, Dong-A University)
Lee, Hyoung-Ju (Department of Applied Biology, Dong-A University)
Moon, Byung-Ju (Department of Applied Biology, Dong-A University)
Lee, Seon-Woo (Department of Applied Biology, Dong-A University)
Publication Information
The Plant Pathology Journal / v.25, no.2, 2009 , pp. 136-141 More about this Journal
Abstract
Marking biocontrol bacteria is an essential step to monitor bacterial behavior in natural environments before application in agricultural ecosystem. In this study, we presented the simple green fluorescent protein (GFP) reporter system driven by the promoter active in Bacillus species for tagging of the biocontrol bacteria. A constitutive promoter P43 from Bacillus subtilis was fused to an enhanced promoterless gfp gene by overlap extension PCR. The GFP expression was demonstrated by the high fluorescence intensity detected in B. subtilis and Escherichia coli transformed with the P43-gfp fusion construct, respectively. The GFP reporter system was further investigated in two bacterial biocontrol strains B. licheniformis and Pseudomonas fluorescens. When the reconstructed plasmid pWH34G was introduced into B. licheniformis, GFP level measured with the fluorescence intensity in B. licheniformis was almost equivalent to that in B. subtilis. However, GFP expression level was extremely low in other biocontrol bacteria P. fluorescens by transposon based stable insertion of the P43-gfp construct into the bacterial chromosome. This study provides information regarding to the efficient biomarker P43-gfp fusion construct for bio-control Bacillus species.
Keywords
Bacillus; biocontrol; GFP; P43;
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Times Cited By Web Of Science : 3  (Related Records In Web of Science)
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1 Chalfie, M., Tu, Y., Euskirchen, G., Ward, W Wand Prasher, D. C. 1994. Green fluorescent protein as a marker for gene expression. Science 263:802-805   DOI   PUBMED
2 Dennis, J. J. and Zylstra, G. J. 1998. Plasposons: modular selfcloning minitransposon derivatives for rapid genetic analysis of Gram-negative bacterial genomes. Appl. Environ. Micro-Biol. 64:2710-2715
3 Fravel, D. R., Connick. Jr. W J. and Lewis, J. A. 1998. Formulation of microorganisms to control plant diseases. In: Formulation of Microbial Pesticides: Beneficial Microorganisms, Nematodes and Seed Treatments, ed. by H. D. Burges, pp 187-202. Kluwer Academic Publishers, Dordrecht, The Netherlands
4 Lee, J. P., Lee, S-W, Kim, C. S., Son, J. H., Song, J. H., Lee, K Y., Kim, H. J., Jung, S. J. and Moon, B. J. 2006. Evaluation of formulations of Bacillus licheniformis for the biological control of tomato gray mold caused by Botrytis cinerea. Biol. Cont. 37:329-337   DOI   ScienceOn
5 Miller, W G. and Lindow, S. E. 1997. An improved GFP cloning cassette designed for prokaryotic transcriptional fusions. Gene 191:149-153   DOI   ScienceOn
6 Park, K., Paul, D., Kim, Y. K., Nam, K W, Lee, Y. K., Choi, H. Wand Lee, S. Y. 2007. Induced systemic resistance by Bacillus vallismortis EXTN-1 suppressed bacterial wilt in tomato caused by Ralstonia solanacearum. Plant Pathol. J. 23:22-25   DOI   ScienceOn
7 Prasher, D. C., Eckenrode, V. K, Ward, W W., Prendergast, F. G. and Cormier, M. J. 1992. Primary structure of the Aequorea victoria green-fluorescent protein. Gene 111:229-233   DOI   ScienceOn
8 Sambrook, J., Fritsch, E. F. and Maniatis, T. 1989. Molecular cloning: A laboratory manual, 2nd ed. Cold Spring Harbor Laboratory, Cold Spring Harbor, N. Y.
9 Jefferson, R.A. 1989. The gus reporter gene system. Nature 342:837-838   DOI   PUBMED   ScienceOn
10 Boyer, H. W. and Roulland-Dussoix, D. 1969. A complementation analysis of the restriction and modification of DNA in Escherichia coli. J. Mol. Biol. 41 :450-472   DOI
11 Prendergast, F. G. and Mann, K. G. 1978. Chemical and physical properties of aequorin and the green fluorescent protein isolated from Aequoreaforskalea. Biochemistry 17:3448-3453   DOI   ScienceOn
12 Cook, R. J. 1993. Making greater use of introduced microorganisms for biological control of plant pathogens. Annu. Rev. Phytopathol. 31 :53-80   DOI   PUBMED   ScienceOn
13 Handelsman, J. and Stabb, E. V 1996. Biocontrol of soilborne plant pathogens. Plant Cell 8:1855-1869   DOI   ScienceOn
14 Burkholder, P. R. and Giles, N. H. 1947. Induced biochemical mutation in Bacillus subtilis. Am. J. Bot. 34:345-348   DOI   ScienceOn
15 Figurski, D. H. and Helinski, D. R. 1979. Replication of an origincontaining derivatives of plasmid RK2 dependent on a plasmid function provided in trans. Proc. Natl. Acad. Sci. USA 76:1648-1652   DOI   ScienceOn
16 Choi, G. J., Kim, J. C., Park, E. J. Choi, Y. H., Jang, K. S., Lim, H. K., Cho, K Y. and Lee, S-W 2006. Biological control activity of two isolates of Pseudomonas fluorescens against rice sheath blight. Plant Pathol. J. 22:289-294   DOI   ScienceOn
17 Cormack, B. P., Valdivia, E. A. and Falkow, S. 1996. FACS-optimized mutants of the green fluorescent protein (GFP). Gene 173:33-38   DOI   ScienceOn
18 Horton, R. M., Hunt, H. D., Ho, S. N., Pullen, J. K. and Pease, L. R. 1989. Engineering hybrid genes without the use ofrestriction enzymes: gene splicing by overlap extension. Gene 77:61-68   DOI   ScienceOn
19 Stepanenko, O. V, Verkhusha, V V, Kumetsova, I. M., Uversky, V N. and Turoverov, K. K. 2008. Fluorescent proteins as biomarkers and biosensors: throwing color lights on molecular and cellular processes. Curr. Protein. Pept. Sci. 9:338-369   DOI   ScienceOn
20 Xue, G. P., Johnson, J. S. and Dalrymple, B. P. 1999. High osmolarity improve the electro-transformation efficiency of the gram-positive Bacillus subtilis and Bacillus licheniformis. J. Microbiol. Methods 34: 183-191   DOI   ScienceOn
21 Wang, P. Z. and Doi, R. H. 1984. Overlapping promoters transcribed by Bacillus subtilis ${\sigma}^{55}$ and ${\sigma}^{37}$ RNA polymerase holoenzymes during growth and stationary phase. J. Biol. Chem. 259:8619-8625   PUBMED
22 Webb, C. D., Decatur, A., Teleman, A. and Losick, R. 1995. Use of green fluorescent protein for visualization of cell-specific gene expression and subcellular protein localization during sporulation in Bacillus subtilis. J. Bacteriol. 177:5906-5911   DOI   PUBMED
23 im, H. J., Lee, S. H., Kim, C. S., Lim, E. K., Choi, K. H., Kong, H. K., Kim, D. W, Lee, S-W and Moon, B. J. 2007. Biological control of strawberry gray mold caused by Botrytis cinerea using Bacillus licheniformis Nl formulation. J. Microbiol. Biotechnol. 17:438-444   PUBMED
24 Kim, G. H., Lim, M. T., Hur, J.-S., Yum, K-J. and Koh, Y. J. 2009. Biological control of tea anthracnose using an antagonistic bacterium of Bacillus subtilis isolated from tea leaves. Plant Pathol. J. 25:99-102   DOI   ScienceOn
25 O'Kane, D. J., Lingle, W L., Wampler, J. E., Legocki, M., Legocki, R. P. and Szalay, A. A. 1988. Visualization ofbioluminescence as a marker of gene expression in Rhizobiuminfected soybean nodules. Plant Mol. Biol. 10:387-399   DOI   ScienceOn
26 Zhang, X-Z., Cui, Z. L., Hong, Q. and Li, S-P. 2005. High-level expression and selection of methyl parathion hydrolase in Bacillus subtilis WB800. Appl. Environ. Microbiol. 71 :4101-4103   DOI   ScienceOn
27 Buell, C. R. and Anderson, A. J. 1993. Expression of the aggA locus of Pseudomonas putida in vitro and in planta as detected by the reporter gene, xylE. Mol. Plant-Microbe Interact. 6:331-340   DOI   PUBMED   ScienceOn
28 Higuchi, R., Krummel, B. and Saiki, R. K. 1988. A general method of in vitro preparation and specific mutagenesis of DNA fragments: study of protein and DNA interactions. Nucleic Acids Res. 16:7351-7367   DOI   ScienceOn
29 Olubajo, B. and Bacon, C. W 2008. Electrotransformation of Bacillus mojavensis with fluorescent protein markers. J. Microbiol. Methods 74:102-105   DOI   ScienceOn
30 Loper, J. E. and Lindow, S. E. 1994. A biological sensor for iron available to bacteria in their habitats on plant surfaces. Appl. Environ. Microbiol. 60: 1934-1941   PUBMED
31 March, J. C., Rao, G. and Bentley, W E. 2003. Biotechnological applications of green fluorescent protein. Appl. Microbiol. Biotechnol. 62:303-315   DOI   ScienceOn
32 Heim, R., Cub itt, A. B. and Tsien, R. Y. 1995. Improved green fluorescence. Nature 373:663-664   PUBMED
33 Mo, Y. Y. and Gross, D. C. 1991. Expression in vitro and during plant pathogenesis ofthe syrB gene required for syringomycin production by Pseudomonas syringae pv. syringae. Mol. Plant-Microbe Interact. 4:28-36   DOI
34 Keane, P. J., Kerr, A. and New, P. B. 1970. Crown gall of stone fruit. II. Identification and nomenclature of Agrobacterium isolates. Aust. J. Biol. Sci. 23:585-595   DOI