Application of Multiplex PCR for Rapid Determination of cryl Gene Profiles of New Bacillus thuringiensis Isolates

The cry1 gene content of a collection of Bacillus thuringiensis strains, which include new isolates from Malaysia and Indonesia, was determined by a multiplex PCR using a set of eight oligonucleotide forward primers specific to cry1Aa, cry1Ab, cry1Ac, cry1Ba, cry1Ca, cry1Da, cry1Ea, and cry1Fa genes, and two reverse primers, one specific to cry1Ab and the other common to the remaining cry1 genes. Two-thirds of the 59 strains screened were cry1 positive and contained one to four different genes. The cry gene profiles correlated well with toxicities of the strains to lepidopteran insects. The method can be used for rapid screening of a large number of new isolates as the total DNA extracted by boiling cells from single colonies can be used directly in the PCR. However, it is not suitable for follow-up monitoring of specific commercial strains after application in the field as the PCR product profiles of these strains could not be differentiated from those of new isolates.