Research Article

Purification of the insecticidal Cry2Ad protein from a Bt-isolated BRC-HZP10 strain and toxin assay to the diamondback moth, Plutella xylostella (L.)

Published: July 13, 2015
Genet. Mol. Res. 14 (3) : 7661-7670 DOI: https://doi.org/10.4238/2015.July.13.11
Cite this Article:
(2015). Purification of the insecticidal Cry2Ad protein from a Bt-isolated BRC-HZP10 strain and toxin assay to the diamondback moth, Plutella xylostella (L.). Genet. Mol. Res. 14(3): gmr5423. https://doi.org/10.4238/2015.July.13.11
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Abstract

The present study aims to characterize the Cry2Ad toxin protein isolated from a Bacillus thuringiensis strain, BRC-HZP10, which have a potential insecticidal activity against larvae of the diamondback moth, Plutella xylostella (L.). The crude Bt toxin proteins were isolated and purified by cation exchange chromatography, then equilibrated with 0.2 M NaOH buffer, pH 4.0, followed by ultraviolet detection at 280 nm and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A refined Cry2Ad toxin protein with 88.34% purity was eventually obtained and used for a bioassay by feeding it to P. xylostella. The results showed conspicuous insecticidal activity towards P. xylostella with 50% lethal concentration of 6.84 μg/mL and 95% confidence interval of 5.77-7.91 mg/mL. At a concentration of 16.38 μg/mL, the intake of Cry2Ad protein significantly shortened the oviposition period and larval developmental duration, but significantly reduced the fecundity and egg hatchability of the population compared to those of control (without treatment with Cry2Ad protein) (P P. xylostella.

The present study aims to characterize the Cry2Ad toxin protein isolated from a Bacillus thuringiensis strain, BRC-HZP10, which have a potential insecticidal activity against larvae of the diamondback moth, Plutella xylostella (L.). The crude Bt toxin proteins were isolated and purified by cation exchange chromatography, then equilibrated with 0.2 M NaOH buffer, pH 4.0, followed by ultraviolet detection at 280 nm and sodium dodecyl sulfate-polyacrylamide gel electrophoresis. A refined Cry2Ad toxin protein with 88.34% purity was eventually obtained and used for a bioassay by feeding it to P. xylostella. The results showed conspicuous insecticidal activity towards P. xylostella with 50% lethal concentration of 6.84 μg/mL and 95% confidence interval of 5.77-7.91 mg/mL. At a concentration of 16.38 μg/mL, the intake of Cry2Ad protein significantly shortened the oviposition period and larval developmental duration, but significantly reduced the fecundity and egg hatchability of the population compared to those of control (without treatment with Cry2Ad protein) (P < 0.05). These results indicate that the Cry2Ad protein plays an effective role in controlling the population of P. xylostella.