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Transformation of signal sequence in Escherichia coli by reporter gene fusion

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Research Paper 01/08/2011
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Transformation of signal sequence in Escherichia coli by reporter gene fusion

Ahmad Humayan Kabir, FM Ali Haydar, M Shibli Abedin
Int. J. Biomol. & Biomed.1( 2), 27-31, August 2011.
Certificate: IJBB 2011 [Generate Certificate]
Key: Erwinia carotovoraGene fusionpHSK34 plasmidSignal sequence

Abstract

Transformation of signal sequence by reporter gene fusion technology was attempted. The promoter containing signal sequences was taken from Erwinia carotovora, a plant pathogenic soil bacterium. Plasmid DNA (Deoxyribonucleic acid) and genomic DNA were cleaved by BamHI and Sau3A1, respectively. Result of gel electrophoresis reveals successful cutting of restriction enzyme in both plasmid and genomic DNA. Best banding was found in 0.75 U/mg DNA of Sau3A1 compared to other concentrations of Sau3A1 used. Bacterial growth was found on LBcmp (Luria-Bertani) plates, since chloramphenicol resistance ability was pre-existed in this plasmid. Highest colony forming unit was observed with ligation ratio (1:5). It was also evident that the cells have been successfully transformed with the plasmid resulting notable growth of bacteria in LBamp plate.

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Albert B, Johnson A, Lewis J, Raff M, Roberts K, P Walter. 2002. Molecular Biology of the Cell (4th Ed.). New York, Garland Science.

Gelbart WM, Griffith, AJF, Lewontin RC, MIller JH, Suzuki D. 2005. An Introduction to Genetic Analysis. (8th Ed.). New York, W.H. Freeman and Company.

Hideakmi A, Tsutoshmi O, Azuhitofu J, Nichki I, Toshiomyio S. 1993. Cloning, sequence analysis and expression in Escherichia coli of a gene encoding an alginate lyase from Pseudomonas sp. OS-ALG-9. Journal of General Microbiology 139, 987-993.

Ian A, Jonathan S, Anna G, Josie H, David LC, Michael AB, Marc V. 2004. Feasibility of Genome-Scale Construction of Promoter::Reporter Gene Fusions for Expression in Caenorhabditis elegans Using a MultiSite Gateway Recombination System. Genome Research 14(10b), 2070–2075.

Gregory JG, Rachel M, Douglas ES. 2006. Tension-dependent DNA cleavage by restriction endonucleases: Two-site enzymes are “switched off” at low force. PNAS 103(31), 11555-11560.

Martinko JM, Madigam MT. 2006. Brock Biology of Microorganisms. (11th Ed.). USA, Pearson Education.

Pannika N, Vanaporn W. 2002. Cloning of the class D β lactamase gene from Burkholderia pseudomallei and studies on its expression in ceftazidime-susceptible and-resistant strains. Journal of Antimicrobial Chemotherapy 50, 445–455.

Peter F, Rudi  L, Gerhild  L, Alfred  P.  2001. Sau3AI, a Monomeric Type II Restriction Endonuclease That  Dimerizes on the DNA and Thereby Induces DNA Loops. The Journal of Biological Chemistry 276, 23581-23588.

Saarilahti HT, Pirhonen M, Karlsson MM, Flego D, Palva DE. 2004. Expression of pehA-bla gene fusions in Erwinia carotovora subsp. carotovora and isolation of regulatory mutants affecting polygalacturonase production. Molecular and General Genetics 234 (1), 81-88.

Shinichi N, Shinichi U, Masamitsu T, Akira H, Masami N, Katsumi A. 2004. Cloning of various promoters for foreign gene expression in Erwinia ananas. Journal of General Plant Pathology 70(1), 69-73.

Sirkka RM, Kyostio CL, George HL. 1991. Erwinia carotovora subsp. carotovora Extracellular Protease: Characterization and Nucleotide Sequence of the Gene. Journal of Bacteriology 173(20), 6537-6546.

Steven DB, George PCS. 2006. Exploitation of a b lactamase reporter gene fusion in the carbapenem antibiotic production operon to study adaptive evolution in Erwinia carotovora. Microbiology 152, 1089–1097.

Tina KVD, Yan W, Michael KH, Maureen D, Mary JGR, Antoni R, Lucia BR, Robert AL. 2001. A genomic approach to gene fusion technology. PNAS 98(5), 2555–2560.

Tiina P, Richard P, Ruusu R, Martin R, Hannu TS. 2002. A putative three-dimensional targeting motif of polygalacturonase (PehA), a protein secreted through the type II (GSP) pathway in Erwinia carotovora. Molecular Microbiology 43(3), 585–596.

Yahua Chen, Fred CT, Theresa MK. 2004. Beta-lactamase Gene Expression in a Penicillin-Resistant Bacillus anthracis Strain. Antimicrobial Agents and Chemotherapy 48(12), 4873–4877.

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