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Isolation and heterologous expression of PHA synthesising genes from Bacillus thuringiensis R1

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Abstract

The polyhydroxyalkanoate biosynthesis gene locus from Bacillus thuringiensis R1 was isolated, cloned and analyzed at the molecular level. We found that a ∼5 kb SacI–ClaI digested fragment of genomic DNA from B. thuringiensis R1 encoding the PHA synthesising genes, conferred PHA producing ability to E. coli. The fragment was sequenced and found to be of 4787 bp with five open reading frames. Sequence alignment with closely related species of Bacillus in the existing database revealed that the ORFs correspond to phaP, phaQ, phaR, phaB and phaC genes. However, E. coli harboring phaP, phaQ, phaR, phaB and phaC locus produced very low PHA. Furthermore, complementation of the locus with phaA from Ralstonia eutropha increased the PHA production in the recombinant E. coli from 3.0% to 24% of cell dry mass. The putative promoter regions and ribosome binding sites were identified for each of the gene. Conserved domains for PHA synthase and aceto-acetyl-coA reductase were also identified. We hence conclude that the PHA operon of Bacillus thuringiensis R1 consists of phaP, phaQ, phaR, phaB, phaC and complementation of the same with phaA is accountable for its high PHA production.

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Acknowledgements

Rohini D. Desetty thanks University Grants Commission, INDIA for the financial support during the work period. We are grateful to Dr. Satoh (Hokkaido University, Japan) for providing us with the primers (P5 and P6) and the plasmid pSRAB. We thank the reviewers for their valuable comments.

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  1. Rohini D. Desetty

    Present address: Department of Biology, York College, City University of New York, New York, NY, 11451, USA

Authors and Affiliations

  1. Plant Tissue Culture Division, National Chemical Laboratory, Pune, 411008, India

    Rohini D. Desetty, Vineet S. Mahajan, B. M. Khan & Shuban K. Rawal

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  1. Rohini D. Desetty
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  2. Vineet S. Mahajan
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  3. B. M. Khan
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Correspondence to Rohini D. Desetty.

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Desetty, R.D., Mahajan, V.S., Khan, B.M. et al. Isolation and heterologous expression of PHA synthesising genes from Bacillus thuringiensis R1. World J Microbiol Biotechnol 24, 1769–1774 (2008). https://doi.org/10.1007/s11274-008-9669-7

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  • DOI: https://doi.org/10.1007/s11274-008-9669-7

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