PCR has been widely used to identify cry-type genes, to determine their distribution, to detect new such genes and to predict insecticidal activities. We describe here a molecular approach to analyze the genetic diversity of B. thuringiensis cry-like genes based on denaturing gradient gel electrophoresis (DGGE). This analysis demonstrated that different B. thuringiensis isolates can be distinguished according to its PCR-DGGE profile of cry-like genes. Identification of the resolvable DNA fragments was easy to accomplish by DNA sequencing, which was confirmed in this work. Importantly, the strategy allowed the identification of unknown B. thuringiensis cry-like sequences present in a single strain that remained cryptic after PCR analysis using degenerate primers. The method developed in this work contributes to the availability of molecular techniques for both B. thuringiensis strains and cry-like genes identification and discovery.