Abstract
Abiotic stresses such as drought, heat and salinity are responsible for decelerating agricultural productivity all over the world and the destructive aftermaths are likely to increase further due to changing global climate. Small heat shock proteins (sHsps) are synthesized ubiquitously and play diverse roles in abiotic stress responses of plants. sHsps are reported to be involved in preventing aggregation, stabilizing non-native proteins. In the present study, Hsp17.9 gene from Prosopis cineraria was cloned, characterized and expression was studied in prokaryotic system. In silico analysis was carried out to gain insights in to protein structure and its interaction with other molecules. Overexpression of the gene in Escherichia coli BL21 (DE3) cells was analyzed on SDS–PAGE, followed by western blotting using Anti His-tag antibody and purified by nickel–nitrilo triacetic acid resin based columns. The single purified band was confirmed by peptide mass fingerprinting. Two dimensional protein analysis of bacterial protein showed the presence of overexpressed protein spot at the desired place. Further, the recombinant E. coli cells carrying pET28a-PcHsp17.9 construct were able to tolerate abiotic stresses better than the cells carrying pET28a vector alone. The cloned PcHsp17.9 gene may be useful to generate abiotic stress tolerant transgenic crops for a climate smart agriculture.
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Abbreviations
- SDS–PAGE:
-
Sodium dodecyl sulfate–polyacrylamide gel electrophoresis
- E. coli :
-
Escherichia coli
- sHsps :
-
Small heat shock proteins
- Ni–NTA:
-
Nickel–nitrilo triacetic acid
- IPTG:
-
Isopropyl-β-D thiogalactoside
- PVDF:
-
Polyvinylidene difluoride
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Acknowledgements
Authors are thankful to the Project Director, NRCPB (National Research Centre on Plant Biotechnology), Delhi, India for providing facilities to carry out research work and Director, IARI (Indian Agricultural Research Institute) for providing National Phytotron Facility, IARI, New Delhi, India. The funds provided by the ICAR (Indian Council of Agricultural Research under NICRA (National Innovations in Climate Resilient Agriculture) project is duly acknowledged
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Experimentation: AK, HV. Writing original draft, data curation, formal analysis: AK, HV, AM. Conceptualization, resources, supervision, review and editing, project administration: JCP. Review and editing: JCP
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Kaur, A., Vishwakarma, H., Maibam, A. et al. Cloning, characterization and in silico studies on abiotic stress responsive Hsp17.9 from Prosopis cineraria. Ind J Plant Physiol. 23, 731–740 (2018). https://doi.org/10.1007/s40502-018-0414-4
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DOI: https://doi.org/10.1007/s40502-018-0414-4