Method for researching influence on generation of microcystic toxin by chloride ions and sulfate radicals by using algae toxin toxin-producing gene

A microcystin and chloride ion technology, applied in the field of molecular biology, can solve the problems of limiting the application of high performance liquid chromatography, difficulty in obtaining standard samples, and low amplification efficiency, achieving high amplification efficiency and accurate detection results , the effect of simple operation

Inactive Publication Date: 2018-01-19
SHANGHAI MUNICIPAL WATER RESOURCES DEV & UTILIZATION NAT ENG CENT +1
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  • Summary
  • Abstract
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  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] Although the above prior art can detect the influence of nitrogen and phosphorus on the generation of algal toxins by detecting the copy number of toxin-producing genes and the concentration of nitrogen and phosphorus, this method needs to use solid phase extraction to pre-process the sample when measuring the concentration of algal toxins. processing, this method takes a long time; this method uses high performance liquid chromatography to determine the concentration of microcystins, and requires technical training for operators. This method relies on microcystin standard samples. More than 80 microcystins have been found. Cystin, most standard samples are difficult to obtain and expensive, which limits the further application of high performance liquid chromatography; this method directly extracts the DNA in the sample, and uses DNA as a template for real-time quantitative PCR, resulting in low amplification efficiency

Method used

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Experimental program
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Embodiment 1

[0040] Cultivate Microcystis aeruginosa with different concentrations of chloride ions, the concentrations of chloride ions in this embodiment are respectively: 0, 50, 100, 200 and 500 mg / liter, and the content of microcystin produced by it is similar to that of the algal toxin synthase gene The correlation before the mcyD copy number is: y=kx+b, where y is the algal toxin content in μg / L, x is the copy number of the algal toxin synthase gene mcyD in copies / mL, and k and b are constant.

[0041] Use different concentrations of chloride ions to act on Microcystis aeruginosa, take 25 milliliters of Microcystis aeruginosa culture fluid, centrifuge with a 10,000 rpm centrifuge for 10 minutes, and remove the supernatant.

[0042] Transfer the algal cells to a 1.5 ml centrifuge tube with phosphate buffered saline, centrifuge at 10,000 rpm for 10 minutes at 4°C, remove the supernatant, collect the algal cells, and store the algal cells in a -80°C refrigerator for subsequent operation...

specific Embodiment approach 2

[0055] Cultivate Microcystis aeruginosa with different concentrations of sulfate radicals. The concentrations of sulfate radicals in this embodiment are respectively: 0, 40, 80, 150 and 300 mg / liter, and the content of microcystin produced by it is comparable to that of the algal toxin synthase gene. The correlation before the mcyD copy number is: y=kx+b, where y is the algal toxin content in μg / L, x is the copy number of the algal toxin synthase gene mcyD in copies / mL, and k and b are constants .

[0056] Use different concentrations of sulfate radicals to act on Microcystis aeruginosa, take 25 ml of Microcystis aeruginosa culture fluid, centrifuge at 10,000 rpm for 10 minutes, and remove the supernatant.

[0057] Transfer the algal cells to a 1.5 ml centrifuge tube with phosphate buffer saline (PBS), centrifuge at 10,000 rpm for 10 minutes at 4°C, remove the supernatant, collect the algal cells, and store the algal cells at -80°C Refrigerator for subsequent operations.

[...

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Abstract

The invention relates to a method for researching influence on generation of microcystic toxin by chloride ions and sulfate radicals by using an algae toxin toxin-producing gene. The chloride ions andsulfate radicals with different concentrations are added in a nutrient solution, the microcystic toxin content and a copy number of the algae toxin toxin-producing gene mcyD are detected, correlationamong the chloride ions and sulfate radicals with different concentrations, the microcystic toxin content and a copy number of the algae toxin toxin-producing gene mcyD is established, and the content of the algae toxin is researched by detecting the copy number of the algae toxin synthetase gene mcyD. The method has the advantages of strong specificity, high sensitivity, short required time, simple operation, and simple required equipment and reagent.

Description

technical field [0001] The invention relates to the field of molecular biology, in particular to a method for studying the influence of chloride ions and sulfate radicals on the production of microcystins by using algae toxin-producing genes. Background technique [0002] Salt tide intrusion occurs all over the world, mostly in coastal areas, especially in estuary areas. Climate change and human activities can increase the frequency and harm of salt tide intrusion. Salt tide intrusion has brought a series of hazards to industry, agriculture and drinking water use. Salt tide intrusion brings a large amount of salt water and ions, changes the photoclimate and mixing characteristics of fresh water, and affects the energy and nutrient cycle in the water. A large amount of salt causes the aggregation and flocculation of suspended matter in the water body, leading to the removal of particulate matter in the water body, increasing the light transmittance of the water body, promot...

Claims

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Application Information

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Patent Type & AuthorityApplications(China)
IPC IPC(8): C12Q1/6851G01N33/53
CPCC12Q1/6851C12Q1/689C12Q2600/158G01N33/53G16B40/00C12Q2545/114C12Q2537/16C12Q2531/113
Inventor陈蕾何义亮姜蕾杨坤刘爽严棋
OwnerSHANGHAI MUNICIPAL WATER RESOURCES DEV & UTILIZATION NAT ENG CENT