Journal of Pure and Applied MicrobiologyVol. 9 No. 3

Cloning and Expression of Variant chiA from Native Serratia marcescens AUDS227

Smita R. Babar*, Sachin A. More and P.U. Krishnaraj

Institute of Agri Biotechnology, University of Agricultural Sciences, Dharwad - 580005, India.

Received on 21 November 2014 and accepted on 08 January 2015

 

ABSTRACT

Serratia marcescens a gram-negative and one of the most effective chitinolytic bacterium used as a model to study the degradation of chitin. The present study was carried out to identify the variant chiA in comparison with the reference Sm141chiA and to study the expression of variant chiA in E. coli and to identify the bioefficacy of variant chiA against the fungal pathogens. Sixty native isolates of S. marcescens were screened to check their chitinolytic activity in comparison with the reference strain Sm141. In order to find out the variability amongst chiA, PCR-RFLP analysis of native chiA positive isolates was done along with the reference Sm141chiA. The variant SmAUDS227chiA identified through PCR-RFLP was cloned in pTZ57R/T and expressed in E. coli BL21 (DE3) by subcloning into prokaryotic expression vector pET32C+ with a molecular mass of ~79.86 kDa. The extracted protein from the cloned SmAUDS227chiA was found effective to inhibit the fungal pathogens viz. R. solani and S. rolfsii.

Keywords : Serratia marcescens, chitin, chitinases, PCR-RFLP, antifungal activity.