A kind of sodium dinitrotoluenesulfonate degrading bacteria and its screening method and application
A technology of sodium dinitrotoluenesulfonate and degrading bacteria, applied in the field of microorganisms, can solve problems such as unreasonable discharge of red water and soil pollution
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Embodiment 1
[0031] Example 1. Screening of sodium dinitrotoluenesulfonate degrading bacteria.
[0032] Screening method:
[0033] 1) Collect the soil polluted by TNT red water near the Gansu military factory, add 5g of soil samples to 50mL inorganic salt medium containing 1 / 10 volume concentration LB medium, and culture on a shaker at 30°C for 48h to obtain cultures;
[0034] 2) Transfer 25mL of the above culture to a new 25mL inorganic salt medium containing 1 / 10 volume concentration LB medium and 10μL substrate (final concentration 20mg / L), and culture on a shaker at 30°C for 24h to obtain a suspension ;
[0035] 3) Take 5mL of the suspension in step 3) and add it to 45mL with a substrate concentration of 50mg / L, containing 20g / L glucose and 200mg / L NH 4 In the inorganic salt medium of Cl, culture on a shaker at 30°C for 7 days.
[0036]4) Subculture is carried out with 7 days as a cycle, and the substrate concentration is gradually increased: 100mg / L, 200mg / L, 300mg / L, 500mg / L, that...
Embodiment 2
[0042] Embodiment 2. The utilization of sodium dinitrotoluenesulfonate degrading bacteria to sodium dinitrotoluenesulfonate.
[0043] The purified single colonies were picked into 5 mL LB medium and cultured overnight at 30°C on a shaker. The seed liquid was taken and streaked on an inorganic salt solid medium plate with a substrate concentration of 500mg / L, cultivated in a 30°C incubator until colonies grew (about 3-4 days), and Escherichia coli JM109 (DE3) was used as a control. as attached figure 2 As shown, the Klebsiella mutans can grow on the inorganic salt solid medium plate with sodium dinitrotoluenesulfonate as the only nitrogen source, while the control group JM109 (DE3) can hardly grow.
Embodiment 3
[0044] Embodiment 3. Degradation efficiency experiment of sodium dinitrotoluenesulfonate by sodium dinitrotoluenesulfonate degrading bacteria.
[0045] Since the additional nitrogen source can promote the degradation of aromatic nitro compounds by microorganisms, we added different concentrations of NH 4 Cl as an additional nitrogen source. Pick a single colony and place it in 50 mL inorganic salt medium with a glucose concentration of 20 g / L and a substrate concentration of 20 mg / L, and culture it on a shaker at 30°C for about 12 hours. Centrifuge at 10000rpm for 10min, wash the bacterial pellet twice with inorganic salt medium, resuspend with a small amount of inorganic salt medium and measure OD 600 value, by OD 600 Value calculated to get transfer amount, transfer to 5mL NH 4 In the inorganic salt medium containing 1g / L glucose and 40mg / L substrate with Cl concentration of 0.5%, 1%, 2%, the initial OD 600 The value was 0.25, and samples were taken after 72 hours of inc...
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