Analysis Of Screening And Identification Of Insecticidal Activity Of Bacillus Thuringiensis From Hengtou Mountain

Agricultural production is plagued by pests. China is a major agricultural country all over the world, but also an agricultural pest disaster-prone country. Agricultural pest has always been one of the huge problems that Chinese agriculture facing. For a long time we use a large number of chemical pesticides to control pests and diseases in China. However, the use of these chemical pesticides has led to several problems, such as environmental pollution and human health effects increasing. Biological control is the best way of solving the series of problems. Bacillus thuringiensis (Bt) is an insecticidal bacterium that has successfully been used as a biopesticide, but traditional Bt strains show some lack such as narrow spectrum of activity and low virulence. So Bt engineering bacteria should build through biological technology for producing need of more efficient and more broad-spectrum. The insecticidal activity of Bacillus thuringiensis mainly comes from the crystal proteins, which formed during sporulation. The mode of action of these toxins and how insects respond to the attack of Cry proteins are a hotspot for study over the world. Understanding the mode of action of these toxins and how insects respond to the attack of Cry proteins will allow the development of changing the cry proteins’ toxicity and active spectrum by using molecular design, for supplying the gene resource of the transgenic plants and microorganism.12Bacillus thuringiensis strains were isolated from200soil samples which collected from Hengtou Mountain, nearby Harbin, in the Chinese province of Heilongjiang. Three strains were selected on the spore-crystal microscopy, insecticidal crystal protein electrophoresis and insecticidal activity, after the biological diversity analysis. There were globular crystal, large and small bipyramidal crystal, cubic crystal types in the3Bt strains. SDS-PAGE showed that strain HTS-S-12expressed130ku protein, strain HTS-S-38and HTS-S-69expressed both130ku and60ku proteins. Spodoptera exigua and Ostrinia furnacalis were sensitive to strain HTS-S-38and HTS-S-69. Strain HTS-S-38contained cry1Aa, cry1Ac, cry1Ia, cry2Aa, cry2Ab, vip3Aa genes, and also contained AiiA, chiB gene. These full-length genes were cloned by using specific primers. Cry2Ab gene was registered in GenBank with the accession number of KJ010821. International Nomenclature Committee of Bt designated it of cry2Ab31. Vip3Aa gene was the same as vip3Aa43. Strain HTS-S-38didn’t show a widely toxicity against many species insect, but it had highly toxic against Lepidoptera, which providing a basic strain for biological pesticide Bt engineering bacteria. Its insecticidal genes supplied more resources for the research of toxic theory, and were meaningful to Bt biological control.