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Biochemical and molecular characterization of δ-endotoxins in Bacillus thuringiensis

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Abstract

The δ-endotoxins (δ-ETX) of four native strains (RT7, RT19, RT25, and RT25), and one reference strain (4L1) of Bacillus thuringiensis were biochemically and molecularly characterized to determine their potential toxic activity against lepidopteran larvae. Crystals of δ-ETX were purified through a two-phase system to determine their morphology, molar mass, solubility, and resistance to proteinases. Toxic activity and cry gene content were also determined. Crystals from native strains exhibited polyhedral, irregular and cuboidal shapes, while those from 4L1 were bipyramidal. Seven proteins with estimated molar mass ≈30–134 kDa were detected as the main components of the native δ-ETX. Only crystals from 4L1, RT24, and RT25 underwend complete solubilization at pH >12.0. Crystals from all strains produced trypsinresistant peptides. None of the cry genes associated with toxicity in lepidopterans (cry1, cry2, cry9) was found in the native strains; however, 4L1 strain harbors cry1 and cry2 genes. Strains RT19 and RT25 caused significant mortality against Trichoplusia ni larvae with partial solubilization at pH 10, strain 4L1 caused 100 % mortality. Toxicity of native strains may come from a novel cry gene.

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Abbreviations

CAPS:

3-cyclohexylamino-1-propanesulfonic acid

δ-ETX(s):

δ-endotoxin(s)

ddH2O:

double-distilled water

dNTPs:

deoxynucleoside triphosphates

DTT:

1,4-dithithreitol

SDS-PAGE:

sodium dodecyl sulfate-polyacrylamide gel electrophoresis

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Authors and Affiliations

  1. Laboratorio de Biotecnología Ambiental, Centro de Biotecnología Genómica — Instituto Politécnico Nacional, 88710, Reynosa, Tamp., México

    N. M. Rosas-García, A. Sánchez-Varela & J. M. Villegas-Mendoza

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  1. N. M. Rosas-García
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  2. A. Sánchez-Varela
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  3. J. M. Villegas-Mendoza
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Correspondence to N. M. Rosas-García.

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Rosas-García, N.M., Sánchez-Varela, A. & Villegas-Mendoza, J.M. Biochemical and molecular characterization of δ-endotoxins in Bacillus thuringiensis . Folia Microbiol 54, 487–492 (2009). https://doi.org/10.1007/s12223-009-0069-7

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