Enrichment culture method of whole-process nitrifying bacteria

A technology of enrichment culture and whole-process nitrification, applied in the field of microorganisms, can solve the problem of relatively few descriptions of enrichment culture methods

Active Publication Date: 2020-09-08
FUDAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

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Problems solved by technology

At present, patent reports on whole-process nitrifying bacteria mainly focus on introducing the characteristics of its enrichment devices, including sand filter enrichment devices (patent docum

Method used

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  • Enrichment culture method of whole-process nitrifying bacteria
  • Enrichment culture method of whole-process nitrifying bacteria
  • Enrichment culture method of whole-process nitrifying bacteria

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0035] Embodiment 1: Enrichment culture of whole process nitrifying bacteria

[0036] Specific steps are as follows:

[0037] 1. Acquisition of enriched culture of nitrifying bacteria throughout the process: water samples are filtered through a sterile 0.22μm filter membrane. Water samples were tap water in the laboratory.

[0038] 2. Place the filter membranes in step 1 in the enrichment medium with low concentration of ammonia, and maintain the pH value of the enrichment medium at 6.9-7.1. The culture time is based on substrate consumption and corresponding concentration of nitrate accumulation as one generation Cultivation cycle. When the substrate concentration was lower than 20% of the initial value, the substrate was added to the initial concentration and the cultivation was continued. After standing in an incubator at 28±1°C and cultivating in the dark for 1 month, take out part of the filter membrane from the enrichment culture flask to extract DNA, and perform quan...

Embodiment 2

[0042] Example 2: Effects of different conditions on the enrichment effect of nitrifying bacteria in the whole process

[0043] Specific steps are as follows:

[0044] 1. Acquisition of enriched culture of nitrifying bacteria throughout the process: water samples are filtered through a sterile 0.22μm filter membrane. Water samples were tap water in the laboratory.

[0045] 2. Place the filter membrane in step 1 in the enrichment medium (pH value range is 6.9-7.1) of different concentrations (nitrogen concentration is 0.1mM, 0.5mM, 1mM and 5mM respectively), at 28±1°C Keep in the incubator and keep away from light. Example 2 detects the enrichment effect of two substrates, ammonia and urea. Ammonia choose ammonia water or NH 4 Cl et al. During the cultivation process, the substrate and product concentrations were regularly detected. When the substrate concentration was lower than 20% of the initial value, an equal amount of filter membrane was taken out from the enrichment...

Embodiment 3

[0047] Embodiment 3: the growth of whole process nitrifying bacteria under the condition of low substrate concentration

[0048] Specific steps are as follows:

[0049] 1. Mix the new sterile filter membrane with the filter membrane that has been enriched with full-range nitrifying bacteria, place them together in a new enrichment medium (pH range is 6.9-7.1), and statically place them in an incubator at 28±1°C. Store in dark place. The culture time is based on the substrate consumption and the corresponding concentration of nitrate accumulation as one generation of culture cycle. Remove the newly added filter when the substrate concentration is less than 20% of the initial value.

[0050] 2. Place the new filter membranes taken out in step 1 respectively in enriched medium with low ammonia or urea concentration (pH range is 6.9-7.1), and culture in an incubator at 28±1°C in the dark. In Example 3, the effects of two substrates were tested, and the substrates were ammonia a...

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Abstract

The invention discloses an enrichment culture method of whole-process nitrifying bacteria. The enrichment culture method comprises the following steps of S1, filtering a water body sample by adoptinga sterile filter membrane; S2, culturing the sterile filter membrane in an enrichment culture medium added with a substrate to obtain a filter membrane subjected to enrichment culture, wherein the substrate comprises ammonia or urea; S3, adopting a new sterile filter membrane as a transfer filter membrane, culturing the transfer filter membrane and the filter membrane subjected to enrichment culture in a new enrichment culture medium added with the substrate, and making the transfer filter membrane contact with the filter membrane subjected to enrichment culture, so that the whole-process nitrifying bacteria are transferred to the transfer filter membrane from the filter membrane subjected to enrichment culture; S4, taking out the transfer filter membrane, and culturing the transfer filtermembrane in a new enrichment culture medium added with the substrate; and S5, continuously transferring the whole-process nitrifying bacteria on the transfer filter membrane for multiple times by adopting the filter membrane, so as to achieve the purpose of enrichment culture of the whole-process nitrifying bacteria. In the culture process, part of the filter membrane is taken out regularly to extract DNA, and quantitative PCR analysis is carried out to confirm the enrichment condition of the whole-process nitrifying bacteria.

Description

technical field [0001] The invention relates to the technical field of microorganisms, in particular to a method for enriching and culturing whole-process nitrifying bacteria. Background technique [0002] Nitrification is a biological process linking nitrogen fixation and denitrification, and plays an important role in the ecosystem nitrogen cycle. Nitrification mainly includes two processes: the ammonium oxidation process in which ammonia is oxidized to nitrite and the nitrification process in which nitrite is oxidized to nitrate. For more than 100 years, it has been believed that nitrification is performed by two different types of microorganisms. First, the ammonia oxidation reaction is completed by ammonia-oxidizing bacteria or ammonia-oxidizing archaea, and then the nitrification reaction is completed by nitrifying bacteria. Two articles in the journal Nature in 2015 [1,2] reported the complete nitrification bacteria (Complete ammonia oxidizer, Comammox) that can ind...

Claims

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Application Information

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IPC IPC(8): C12N1/02C12Q1/6851
CPCC12N1/02C12Q1/6851C12Q2531/113Y02W10/10
Inventor 全哲学蒋秋悦夏飞戴红
Owner FUDAN UNIV
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