Screening Of Biological Control Bacillus And Isolation And Purification And Identification Of Antibiotic Substances

Crop disease is a global problem, it causes a huge economic losses. Crop disease is mainly caused by fungus. Biological control of crop disease is the techniques and methods of use beneficial microbes or it’s metabolites for effective prevention and control of crop diseases. Bacillus have lots of advantages, such as high resistance of harsh environment, widely distribute, quick growth characteristics and so on. Bacillus draw more and more attention of researchers in the area of biological control.The research is begin with screening of Bacillus from Heilongjiang Province in Liangshui National Nature Reserve Forest soil and repeat purification. Use plant pathogenic fungus as target strain to screen biological control Bacillus. Then preliminary get dozens of strains of Bacillus. After two times of screening, Bacillus strains with stable antibacterial activity were got. Ultimately three strains of biological control Bacillus with high antibacterial activity were selected and preserved.From the aspects of morphological identification and physiological and biochemical identification, 16 Sr DNA sequence determination to identify three strains of biological control Bacillus. After serious identify, these three strains are Bacillus pumilus F1, Paenibacillus polymyxa F17 and Bacillus subtilis H1.The antagonistic mechanism of biological control strains screened were investigated, find the fermentation metabolites of these three strains of biological control Bacillus have obvious inhibitory effects on plant pathogenic fungus. The research initially determine that biological control Bacillus inhibit the growth of plant pathogenic fungus through secreting antibiotic substances. Different biological control Bacillus has different degree of inhibition on the same plant pathogenic fungus, and the same biological control Bacillus also have different inhibitory effect on the different plant pathogenic fungus. And after high temperature treatment, biological control Bacillus strain F1 and H1 has obvious antibacterial activity, while biological control Bacillus strain F17 lose the antibacterial activity after treatment.The crude protein of Bacillus pumilus F1 was precipitated by using saturation concentration of 60%. After that, using ion exchange chromatography and gel filtration chromatography. The purified fractions were tested of antibacterial activity. The purified protein were tested by sodium dodecyl sulphate polyacrylamide gel electrophoresis(SDS-PAGE). It was found that the antibiotic protein molecular weight is about 36.0 k Da. The antibiotic protein still has obvious antibacterial activity after high temperature treatment for 30 min.In this paper, Bacillus with the function of inhibiting the growth of plant pathogenic fungus was obtained. Antibiotic protein has been isolated and purified, which lays the foundation for future research.