A Burkholderia Bacteria For Biological Control Of Genetically Modified (gm), And Growth Conditions

Burkholderia is a new biopesticide that can resist many plant fungal diseases and can produce many antibiotics. We isolate a Burkholderia KDMD from earth. We design primers according to Pseudomonas aeruginosa hcnABC sequence and amplify hcnABC 2.1kb fragment and 1.6 kb fragment. Later use BamHI and Pstl digest 2.1kb or Pucp26 plasmid, and ligate 2.1kb to Pucp26. Because Pucp26 has LacZ gene, we can select positive colony by blue white method. Check up plasmid by digestion. Then use Hindlll and Pstl to digest 1.6kb and transformed Pucp26, then ligate 1.6kb to transformed Pucp26 and form whole hcnABC gene. We incubate Burkholderia KDMD at different temperature such as 25 ℃, 30℃, 37℃, 42℃ and different pH such as pH5.5-8.5 in LB medium and measure OD₆₀₀ every 2 hours, then draw growth curve and make sure 37℃ is the best growth temperature and pH6.0 is the best growth pH. Then we choose M9 as basic medium. We choose 0.1% yeast extracts as nitrogen source and choose sucrose, glucose, fructose, lactose, starch as carbon source or 1% glucose as carbon source and yeast extracts, peptone, NH₄Cl, NaNO₃ as nitrogen source. We measure OD₆₀₀ every 2 hours, and the draw growth curve and confirm sucrose is the best carbon source and yeast extracts is best nitrogen source. We calculate Burkholderia KDMD culture antagonistic effect to 4 plant pathogen fungi by filter paper method in PDA medium. Then we find antagonistic effect from big to small is Pythium solani, Fusarium oxysporium f.sp.niveum, Gibberella nicotiancola and Gibberella saubinetii. We find Burkholderia KDMD can produce little siderophores in CAS medium and Burkholderia KDMD do not produce HCN by picric acid method. We find Burkholderia KDMD can restrain Gibberella saubinetii in pot experiment.