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A method for detecting polysaccharide bacteria in activated sludge of sewage treatment using sybr GREEN I single-staining technique

A technology for sewage treatment and activated sludge, applied in biochemical equipment and methods, measurement devices, microbe determination/inspection, etc., can solve the problems of complex experimental operation, loss of fluorescence, and no research reports, etc., and achieve simple experimental operation , reliable results, simple operation

Active Publication Date: 2021-10-15
BEIJING UNIV OF TECH
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Problems solved by technology

[0007] (3) The traditional method mainly uses double dyes to detect bacteria, that is, FITC is used to stain the whole bacteria, and CY5 or CY3 is used to stain the target bacteria. The experimental operation is more complicated; Fluorescence loss, so fluorescent probes need to be strictly protected from light during use and storage; and a good hybridization environment needs to be created during probe hybridization in order to better enter the bacteria for DNA double-strand binding
[0008] Therefore, the present invention takes the activated sludge microorganisms in the sewage treatment system, adopts SYBR GREEN I single-staining combined flow cytometry technology, and utilizes the bacterial volume and intracellular particle complexity to analyze the percentage content of polysaccharide bacteria in the microbial community. Related Research Reports

Method used

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  • A method for detecting polysaccharide bacteria in activated sludge of sewage treatment using sybr GREEN I single-staining technique
  • A method for detecting polysaccharide bacteria in activated sludge of sewage treatment using sybr GREEN I single-staining technique
  • A method for detecting polysaccharide bacteria in activated sludge of sewage treatment using sybr GREEN I single-staining technique

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Experimental program
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Embodiment Construction

[0029] (1) Sludge microbial immobilization

[0030] a. Take 300 μL of mud sample in a 2ml centrifuge tube, centrifuge at 10000r / min for 3 minutes,

[0031] Discard the supernatant and add 300 μL of PBS buffer;

[0032] b. Add 1.2ml of paraformaldehyde with a mass percentage concentration of 4% to the centrifuge tube and solidify at 4°C

[0033] Set 3h;

[0034] (2) Cleaning and dilution after fixation

[0035] a. Take out the centrifuge tube from 4°C and centrifuge at 14000r / min for 2min30s;

[0036] b. remove the supernatant;

[0037] c. Add 1.2ml of 1×PBS phosphate buffer solution to the tube, centrifuge to remove the supernatant; repeat this procedure

[0038] step twice;

[0039] d. Add 300 μL of 1×PBS phosphate buffer solution to the tube;

[0040] (3) Ultrasonic dispersion and ethanol dehydration

[0041] aUse an ultrasonic breaker, the ultrasonic power is 80W, stop for 5s after every 30s of ultrasonication, for a total of 4min30s;

[0042] b Divide the ultrason...

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Abstract

The invention discloses a method for detecting polysaccharide bacteria in sewage treatment activated sludge by using SYBR GREEN I single-staining technology, which belongs to the technical field of sewage biological treatment. According to the physical properties of polysaccharide bacteria combined with SYBR GREEN I single staining technology, the present invention detects its content in the sewage treatment system. The morphological characteristics of the whole bacteria were detected by flow cytometry, and the percentage of glycan bacteria in the whole bacteria was analyzed according to the size of the bacteria and the complexity of intracellular particles, and then the relative quantification of the glycan bacteria was done. The invention provides an economical, convenient and reliable quantitative method for polysaccharide bacteria in activated sludge in a sewage treatment system, and provides a theoretical basis for regulating and controlling biological phosphorus removal conditions in actual sewage treatment plants.

Description

technical field [0001] The invention relates to a method for detecting polysaccharide bacteria in sewage treatment activated sludge by using SYBR GREEN I single staining technology, belongs to the technical field of sewage biological treatment, and is used for quantitative analysis of polysaccharide bacteria in sewage biological treatment systems. Background technique [0002] Water eutrophication has become one of the important environmental problems faced by my country's sustainable development, and nitrogen and phosphorus are the main factors that produce water eutrophication. At present, the more economical and effective phosphorus removal method is the enhanced biological phosphorus removal method based on the excessive phosphorus uptake of phosphorus accumulating bacteria. In the EBPR system, in addition to phosphorus-accumulating bacteria, there is another type of microorganisms—polysaccharide bacteria, which can also absorb carbon sources in an anaerobic environment ...

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): G01N15/14C12Q1/04
Inventor 曾薇孟庆安王保贵彭永臻
Owner BEIJING UNIV OF TECH
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