Ninety-two Bacillus thuringiensis isolates were screened from 683 soil samples collected from tropical and semitropical primeval forests in Yunnan and Hainan provinces of China. Several shapes of crystals,including bipyramidal,square,ovoid,spherical,and amorphous,were observed in the B. thuringiensis isolates. Twenty-six pairs of primers were used to identify 31 holotype cry genes at primary rank of the B. thuringiensis cry gene nomenclature system. The cry gene-types of 92 B. thuringiensis isolates and 33 B. thuringiensis isolates screened from Northeast region of China were identified by PCR-RFLP and SDS-PAGE methods. Fifty-eight isolates harbored cry1 genes,32 isolates cry2 genes,12 isolates cry8 genes,3 isolates cry9 genes,12 isolates cry11 genes,and 13 isolates cry30 genes. Of the tested isolates,42 produced no reaction product with 26 pairs of primers and also exhibited no toxicity against 8 insect species tested. The isolate Z2-34 harbored a novel cry30 gene,exhibited insecticidal activity against Aedes albopictus of Dipterans. The accession number of the novel genes in this study is AY916046. Isolation and identification of B. thuringiensis and cry gene are important for investigating the diversity of B. thuringiensis resources and cloning new cry gene. Ninety-two Bacillus thuringiensis isolates were screened from 683 soil samples collected from tropical and semitropical primeval forests in Yunnan and Hainan provinces of China. Several shapes of crystals, including bipyramidal, square, ovoid, spherical, and amorphous, were observed in the B. Thuringiensis isolates. Twenty-six pairs of primers were used to identify 31 holotype cry genes at primary rank of the B. thuringiensis cry gene nomenclature system. The cry gene-types of 92 B. thuringiensis isolates and 33 B. thuringiensis isolates screened from Northeast Fifty-eight isolates harbored cry1 genes, 32 isolates cry2 genes, 12 isolates cry8 genes, 3 isolates cry9 genes, 12 isolates cry11 genes, and 13 isolates cry30 genes. Of the tested isolates 42 produced no reaction product with 26 pairs of primers and also exhibited no toxicity against 8 insect species tested. The isolate Z2-34 harbored a novel cry30 gene, extracted insecticid al activity against Aedes albopictus of Dipterans. The accession number of the novel genes in this study is AY916046. Isolation and identification of B. thuringiensis and cry genes are important for investigating the diversity of B. thuringiensis resources and cloning new cry gene.