African Journal of Biotechnology Vol. 6 (15), pp. 1821-1827, 6 August 2007   

ISSN 1684–5315 © 2007 Academic Journals        

Full Length Research Paper

Characterisation of Bacillus thuringiensis kurstaki strains by toxicity, plasmid profiles and numerical analysis of their cryIA genes

S. Kashyap ¹ and D. V. Amla ²*

¹National Bureau of Agriculturally Important Microorganisms (ICAR) Kushmaur Katholi, Maunath Bhangan, 275101 Uttar Pradesh, India.

²Molecular Biology and Genetic Engineering Division, National Botanical Research Institute, Rana Pratap Marg, P. B. # 436, Lucknow 226001, Uttar Pradesh, India.

*Corresponding author. E-mail: sudhanshukshyp@yahoo.com.

Abbreviations: ICP, Insecticidal Crystal Protein; H.a., Helicoverpa armigera; S.p., Spodoptera littura; Bt, Bacillus thuringiensis; Btk, Bacillus thuringiensis kurstaki; RH, relative humidity; BSA, bovine serum albumin; kDa, kilodalton; SDS-PAGE, Sodium Dodeosyl Sulfate Poly-acrilamide  Gel Electrophoresis; BME, ß-Mercaptoethanol;  and DTT, Dithiothreitol.

Accepted 25 April, 2007

In the present comparative study, four Bacillus thuringiensis kurstaki strains (HD1, dipel, HD73, HD1dipel) were characterized by investigating their total plasmid profiling; cryIA genes profiling and toxicity towards local isolates of agricultural insects Helicoverpa armigera and Spodoptera litura. Result showed that LC₅₀ for S. litura were 0.11 μg of HD73, 0.027 μg of HD1, 0.20 μg of dipel and 0.018 μg of HD1 dipel, while LC₅₀ for H. armigera insect were 0.04 μg of HD73, 0.031 μg of HD1, 0.011 μg of dipel and 0.008 μg of HD1dipel. The native plasmid number and type varied from 4 - 8 among these strains and HindIII restriction showed 0.4 to 8.0 kb size fragments. The numerical classification of the cryIA gene profiles showed two distinct clusters at 48% similarity level. Cluster one of 80% similarity comprises of two strains HD73 and dipel, while cluster two of 65% consisted of HD1 and HD1dipel strains. Southern analysis of restricted plasmid combined with cryIA profile can provide an effective approach for investigation of taxonomic relationship within Bacillus thuringiensis kurstaki strains.

Key words: Bacillus thuringiensis kurstaki, Helicoverpa armigera, Spodoptera litura, δ-endotoxin, plasmids, Southern hybridization.