Heekyu Choi

Dual Insecticidal Activity of Spodoptera-Toxic Bacillus thuringiensis Strain Transformed with Lepidopteran-Specific Cry Toxin

Abstract The E. coli-B. thuringiensis shuttle vector for expression of cry1Ac , pHT1K-1Ac plasmid was introduced into acrystalliferous B. thuringiensis Cry − B and Spodoptera toxic STB-3 strain. The presence of a recombinant plasmid in transformants after electroporation was confirmed by PCR. The 1K-1Ac/Cry − B(Cry − B transformant) and 1K-1Ac/STB-3 (STB-3 transformant) produced bipyramidal-shaped parasporal inclusion that was 130 kDa in size as like B. thuringiensis subsp. kurstaki HD-73. In P. xylostella bioassay, these transformants showed significantly high toxicity than the wild-type recipients and further, in case of B. thuringiensis STB-3 transformant still had original Spodoptera toxicity. These results suggested that the pHT1K could be successfully applied for generating individual B. thuringiensis strains that produce various combinations of insecticidal proteins to expand their host spectrum and enhance insecticidal activity.

Identification of a Novel Crystal Protein Gene from Bacillus thuringiensis Isolated in Korea

To identify novel cry1-type crystal protein genes, 100 Bacillus thuringiensis (Bt) isolates were selected on the basis of their toxicity against lepi- dopteran insect larvae. For rapid search of a large quantity of novel genes simultaneously, Bt isolates were randomly divided into 2 groups including 50 isolates each. About 2.4 kb PCR fragments were amplified from each group using universal oligonu- cleotide primers, ATG1-F and N400-R, designed to probe the toxic fragment regions of all known and possible cry1-type genes. Restriction fragment length polymorphism (RFLP) analysis revealed 3 distinct patterns in each group. One of them, A32 clone showed 87% and 91% of nucleotide sequence and deduced amino acid sequence similarities with the cry1Ea gene. Interestingly, nucleotide sequence ana- lysis of A32 cry gene suggested that this gene might have resulted from nucleotide rearrangement between cry1Ac and cry1Ea genes.