| Schisandra chinensis is one of the important native drugs in the northeast of China. The harm to artificial field and artificial raising semi-wild field of Schisandra chinensis caused by the drift of spraying herbicide in the surrounding plots has become increasingly serious. Among them, two main pesticides are clomazone and aerofloat sprayed in soybean fields, and2,4-D butylate sprayed in corn fields. Using these two kinds of pesticides respectively as the sole carbon source in this study, the strains that could degradate these two kinds of pesticides efficiently were cultivated and screened, and the biological characteristics of the strains as well as the distributional characteristics of degrading enzymes was analysed.1. The bacterias that could degradate corresponding pesticide were isolated and purified from the soil harmed by the drift of clomazone,2,4-D butylate, according to pesticide concentrations in soil and the concentrations of pesticides used as the sole carbon source, nine strains that could degradate clomazone or2,4-D butylate were isolated and screened. Based on the biological measurement, the concentrations of pesticides were measured, and the degradation rates of every strain were analysed, among them, the degradation rate of strain G8that could degradate clomazone efficiently was55.6%, the degradation rate of strain D8that could degradate2,4-D butylate pesticide efficiently was53.2%. Mix of various bacterias could contribute significantly to the degradation of clomazone or2,4-D butylate.2. The strains G8and D8were observed for their morphology in general and electron microscopy. Based on this, further extraction of DNA from strains that could degradate pesticides was carried on by CTAB method, and the bacterial ribosome specific fragment was amplified with general primers for16S rDNA, then cloned and sequenced. By sequence analysis, combined with morphological characteristics, strain G8that could degradate clomazone was identified as Methylobacillus sp., strain D8that could degradate2,4-D butylate pesticide was identified as Rhodocyclus sp..3. In conditions of pot culture, seedlings of Schisandra chinensis harmed by clomazone and2,4-D butylate were inoculated with degrading bacterias, according to analysis of chlorophyll content. After the harm is caused by clomazone and2,4-D butylate, the chlorophyll content in leaves of Schisandrae chinensis decreased, and the chlorophyll content rises with inoculation of degrading bacterias, which was presumably associated with the degradation of clomazone or2,4-D butylate caused by degradating bactetias.4. Using the method of SDS, total DNA was extracted from soil bacteria in soil. Using this total DNA as the template, PCR failed to amplify specific fragment, it maybe be the reason that impurities affect the activity of DNA polymerase. After ten times dilution of the DNA, the target product was amplified efficiently. Presumably by dilution process, the impurities content in the nucleic acid reduced, so their impact on the activity of DNA polymerase reduced too. By this technical system, using GC clamp primers, bacterial group in the soil could be effectively amplified, which laid a foundation for using denaturing gradient gel electrophoresis technical to analyze the influence for bacterial communities in soil after adding the degradation bacteria.5. The most suitable growing conditions of the efficient degradating bacterias of clomazone and2,4-D butylate are established, among them, the optimal pH is7.0. The most suitable growth temperature of Clomazone-efficient degradating bacterial strain is30℃, and the most suitable growth temperature of2,4-D butylate-efficient degradating bacterial strain is35℃. The best speed to culture clomazone,2,4-D butylate efficient degradating bacteria is200r/min. The most suitable ventilation of clomazone-degrading bacteria is100mL/250mL, and the most suitable ventilation of2,4-D butylate-degradating bacteria is120mL/250mL. The optimum pesticide concentration to culture clomazone efficient degradating bacteria is100mg/L, and the optimum pesticide concentration to culture2,4-D butylate-degrading bacteria is500mg/L.6. To collect the bacteria body and the supernatant of strains G8and D8, and prepare the broken liquid of the bacteria body, and add the clomazone and2,4-D butylate to the supematant and the broken liquid of the bacteria body, then insulate them for8hours, after that, the contents of the clomazone and2,4-D butylate are analysed. Based on the result that the contents of the clomazone and2,4-D butylate are cut down, it is conjectured that degarding enzyme might be the extracellular enzymes which exist in the supematant. |
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