A kind of heterotrophic nitrifying aerobic denitrifying Citrobacter and its application
A technology of Freund's citric acid and bacillus, applied in the field of applied microorganisms, can solve the problems of ammonia nitrogen cannot be processed in time, shorten the hydraulic retention time, and long hydraulic retention time, and achieve good biocatalytic function, wide temperature adaptability and growth adaptability strong effect
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Embodiment 1
[0028] Enrichment culture and isolation of embodiment 1 Citrobacter freundii S06
[0029] The activated sludge was taken from the treatment pond of Shanghai Pudong Zhuyuan No. 1 Sewage Treatment Plant, and 5mL of mud-water mixture was inoculated into 45mL of sterilized NRM enrichment medium (the formula of NRM medium was: (NH 4 ) 2 SO 4 2.0g / L, sodium citrate 20.0g / L, MgSO 4 ·7H 2 O 1g / L, 1.0mM phosphate buffer saline K-PBS, pH 6.8. ); 30°C, 200rpm, cultured on a constant temperature shaker for 24h; then the enriched seed solution was serially diluted, and coated onto an NRM solid culture plate (NRM, containing 2% agar); cultivated in a constant temperature incubator at 30°C After 24 hours, the grown microbial clones were cultured separately, inoculated into liquid bromothymol blue (BTB) medium, and cultured in a constant temperature incubator at 30° C. for 24 hours. The formula of BTB medium is (per liter): KNO 3 1g, sodium succinate 8.5g, MgSO 4 ·7H 2 O 1g, CaCl 2...
Embodiment 2
[0030]Identification and Genetic Tree Analysis of Example 2 Bacterial Strains
[0031] Collect 2ml of fresh strain culture solution, centrifuge at 10,000g for 2min, use a genomic DNA extraction kit, such as the kit from Axygen, to extract the genomic DNA of S06, dissolve and dilute to 100mg / L, and use this as a template to amplify 16SrDNA The primers used were 16S-F and 16S-R, and the sequences were 16S-F: AGAGTTTGATCCTGGCTCA, and 16S-R: GGTTACCTTGTTACGACTT. The PCR reaction mixture was prepared as follows: template DNA 1 μL, PCR Taq mix 25 μL, upstream and downstream primers 1.0 μL (20 μM), add ddH 2 0 to 50 μL. The PCR program was run as follows: 94°C for 5 min, 94°C for 30 s, 53°C for 1 min, 72°C for 2 min, 30 cycles, 72°C for 5 min, 4°C for 1 min. The PCR product was sequenced by a professional biological company, such as Shanghai Sangon Biotechnology Co., Ltd. The obtained sequence was analyzed by Blasm using a network tool, and the genetic tree analysis was completed b...
Embodiment 3
[0033] The cultivation and biological deamination analysis of embodiment 3Citrobacter freundii S06
[0034] Inoculate the monoclonal colony on the LB plate into 50ml of LB liquid medium, 30°C, 200rpm, shake and culture in a 250ml shake flask for 24h, take 3% (v / v) culture solution and centrifuge, 3,000g, 2min, to Wash with 1ml of NRM medium, then inoculate into 100ml of NRM liquid medium, shake and culture in a 500ml shaker flask for 72h at 30°C, 200rpm, regularly sample and measure ammonia nitrogen (NH 3 -N), nitrate (NO 3 -N) and nitrite (NO 2 -N) concentration, by analyzing the residual amount of total nitrogen (TN) to measure the removal rate of the strain to inorganic nitrogen, and thus to judge the heterotrophic nitrification and aerobic denitrification performance of the strain. Depend on figure 2 It can be seen that the biological denitrification performance of the isolated strain S06 in the synthetic medium is very high, and it has a good denitrification performan...
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