CONSTRUCTION OF AN EXPRESSION SHUTTLE VECTOR FOR ESCHERICHIA COLI AND BACILLUS SUBTILIS

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RASHIDI MOHAMMAD REZA; MOGHBELI MAJID

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Journal: JOURNAL OF MICROBIAL WORLD Year:2013 | Volume:6 | Issue:3 (16) Page(s): 188-197

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Title

CONSTRUCTION OF AN EXPRESSION SHUTTLE VECTOR FOR ESCHERICHIA COLI AND BACILLUS SUBTILIS

Author(s)

RASHIDI MOHAMMAD REZA | MOGHBELI MAJID | Issue Writer Certificate

Keywords

ESCHERICHIA COLIQ1

BACILLUS SUBTILISQ2

CLONINGQ2

SHUTTLE VECTORQ2

Abstract

Background and Objectives: BACILLUS SUBTILIS is considered as a suitable host for gene CLONING and protein expression due to non-pathogenicity and ability to secrete high amounts of proteins. However, transformation efficiency of ligated plasmid DNA into competent cells of B. subtilis is low, as compared to that into CaCh shocked cells of ESCHERICHIA COLI. Therefore, CLONING the target in E. coli using a SHUTTLE VECTOR before transfer into B. subtilis by a high amount of hybrid vector is more efficient. The aim of this study is construction a SHUTTLE VECTOR using p W980 and pET-I6b plasmids for gene CLONING and expression in B. subtilis.Materials and Methods: p WB980 plasmid was isolated from its host using alkaline lysis method. It was undergone a double digestion to obtain the segment of interest using EcoRI and SnaBI enzymes. Then a special fragment of pET-I6b plasmid was also amplified by PCR and was double digested. Ligation reaction was performed between these two segments and then it was transferred to E. coli TOPIO. Following screening the cells containing SHUTTLE VECTOR, plasmid was extracted and was transferred to B. subtilis WB600 by an effective method.Results: The resulting SHUTTLE VECTOR replicated easily in both hosts. It showed good stability in B. subtilis, which is helpful for its maintenance in its host.Conclusion: In this study, the resulting SHUTTLE VECTOR-pMRI2-had 8 unique CLONING site in its polylinker and a suitable size (5.4 kb), which make it possible to simply gene CLONING into it. This is the first report of construction of an expression SHUTTLE VECTOR for ESCHERICHIA COLI and BACILLUS SUBTILIS in Iran.

Cites

EVALUATION OF THE EFFECT OF SMOKE OF ANBAR NASARA (DONKEY DUNG) EXTRACT ON STAPHILOCOCCUS AUREUS AND BACILUS SUBTILIS

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APA: Copy

RASHIDI, MOHAMMAD REZA, & MOGHBELI, MAJID. (2013). CONSTRUCTION OF AN EXPRESSION SHUTTLE VECTOR FOR ESCHERICHIA COLI AND BACILLUS SUBTILIS. JOURNAL OF MICROBIAL WORLD, 6(3 (16)), 188-197. SID. https://sid.ir/paper/189398/en

Vancouver: Copy

RASHIDI MOHAMMAD REZA, MOGHBELI MAJID. CONSTRUCTION OF AN EXPRESSION SHUTTLE VECTOR FOR ESCHERICHIA COLI AND BACILLUS SUBTILIS. JOURNAL OF MICROBIAL WORLD[Internet]. 2013;6(3 (16)):188-197. Available from: https://sid.ir/paper/189398/en

IEEE: Copy

MOHAMMAD REZA RASHIDI, and MAJID MOGHBELI, “CONSTRUCTION OF AN EXPRESSION SHUTTLE VECTOR FOR ESCHERICHIA COLI AND BACILLUS SUBTILIS,” JOURNAL OF MICROBIAL WORLD, vol. 6, no. 3 (16), pp. 188–197, 2013, [Online]. Available: https://sid.ir/paper/189398/en

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