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In agriculture, chitinolytic microorganisms have been widely used for biocontrol of fungal phytopathogens as cell wall of most fungal pathogens contains chitin to varying extent. Paenibacillus species are widely distributed in nature and have been reported for their biocontrol potential against bacteria and fungi owing to the production of cell wall degrading enzymes such as β-1,3-glucanases, cellulases, chitinases and proteases. This study indicated that plant growth promoting rhizobacterium P. polymyxa has higher chitinase activity compared to Bacillus thuringiensis which is the most widely used biopesticide and known primarily for its chitinase. A chitinase gene (chi45) from P. polymyxa was cloned, sequenced, and expressed in Escherichia coli. The gene consisted of an open reading frame of 1,248 nucleotides encoding a protein of 416 amino acids with a calculated molecular weight of 45,760 Da. Zymogram analysis indicated that the purified enzyme revealed the presence of the 45-kD chitinase. The purified chitinase was shown to inhibit the growth of Rhizoctonia solani, plant pathogenic fungus. Characterization of novel chitinase from P. polymyxa suggested that this enzyme is applicable for various industrial applications, including its use as biopesticide.