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Scapharca subcrenata catalase as novel marine biological pollution detection marker

A technology for catalase and marine pollution in clams, which is applied to the determination/inspection of oxidoreductases, microorganisms, biochemical equipment and methods, etc., to achieve the effects of reducing the accumulation of bases, having strong specificity and improving the degree of binding.

Inactive Publication Date: 2018-11-16
ZHEJIANG OCEAN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

However, the detection of marine pollution with catalase has not yet been reported.

Method used

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Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0028] The clam catalase gene, the clam catalase gene is a cloned gene; the clam catalase gene is cloned and obtained by a gene cloning technique.

[0029] The concrete steps of the cloning of clam catalase gene are:

[0030] 1) Extraction of total RNA: extract total RNA from each tissue using cockles as material;

[0031] 2) Synthesis of the first strand of cDNA: using the total RNA sample obtained in step 1 as a template, use the M-MLV reverse transcription kit to perform reverse transcription synthesis of cDNA to obtain the first strand of cDNA;

[0032] 3) PCR amplification of the core fragment of the clam catalase gene: According to the sequence of the mussel clam catalase gene in the NCBI database, the conserved region of the amino acid sequence was analyzed, and the primer5.0 software was used to design degenerate primers to amplify the clam The core sequence of catalase is subjected to PCR amplification reactions to obtain product fragments of different lengths, and t...

Embodiment 2

[0046] The clam catalase gene, the clam catalase gene is a cloned gene; the clam catalase gene is cloned and obtained by a gene cloning technique. The concrete steps of the cloning of clam catalase gene are:

[0047] 1) Extraction of total RNA: extract total RNA from each tissue using cockles as material;

[0048] Take a RNase-free 2ml EP tube, add 500ul Trizol to the tip of the pipette used, pick up the brain tissue sample with tweezers soaked in alcohol and dissolve it in the Trizol liquid, grind it with an electric homogenizer until the tissue is completely dissolved in the liquid, and fill up the Trizol Centrifuge at 1mL, 4°C, 12000rpm, 10min, take the supernatant and put it into a 1.5mL EP tube. Impurities cannot be absorbed, then add 200ul of chloroform, shake vigorously, let stand at room temperature for 5 minutes, centrifuge at 4°C, 12000rpm, 15min , take the uppermost liquid of the three layers into a new 1.5mL EP tube, add 500ul of isopropanol, room temperature for ...

Embodiment 3

[0128] Using 15-day-old cockles as biological samples for monitoring water pollution, the cockles were divided into two groups, the non-polluted control group and the water area to be tested group, and the cockles were raised in the non-polluted water body and the water area to be tested, respectively, with 3 in each group. After feeding for 7 days, 3 samples were taken in each parallel for measurement, and 3x3=9 data were measured in each group, and the results of each group of cockles were obtained after mathematical statistics of the 9 data in each group.

[0129] The relative expression levels of clam catalase in blood cells and gills were measured using the steps in Example 2. Test results: the relative expression values ​​of the control group in blood cells and gills were 2.37 and 1.96, and the relative expression values ​​of the test group in digestive glands were 1.95 and 1.34. The results showed that the relative expression values ​​of the control group were greater th...

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Abstract

The invention discloses a scapharca subcrenata catalase gene. The scapharca subcrenata catalase gene is a clone gene which is prepared by cloning a core segment of the scapharca subcrenata catalase gene by using a gene cloning technology. The invention further discloses a novel marine biological pollution detection marker. The marine biological pollution detection marker is characterized in that scapharca subcrenata is taken as a biological sample for detecting marine pollution; the scapharca subcrenata catalase is taken as a detection marker for the marine pollution; the relative expression quantity of the scapharca subcrenata catalase gene is the highest in blood corpuscle and cheekbones of the scapharca subcrenata; a PCR (Polymerase Chain Reaction) reaction system is adopted in order toensure the specificity of an amplification reaction, and increase PCR amplification efficiency. The detection marker provided by the invention has high specificity, a sensitive biological reaction isperformed at a reaction endpoint by directly using a biological in vivo target cell or a target molecule at the molecular level, and exposure and toxicity effect of marine pollutant can be alarmed.

Description

technical field [0001] The invention relates to the technical field of molecular biology, in particular to clam catalase, a novel marine biological pollution detection marker. technical background [0002] The sea area of ​​the East my country Sea in China is in the transition zone between the sea and the land, where most of the wastewater discharged from land-based industrial and agricultural production and marine industry development gathers. Among them, pollutants such as heavy metals and petroleum (BAP) are easily combined to the surface of suspended particles or colloids through adsorption and other effects, and are precipitated and enriched, which will have a greater impact on benthic organisms. Studies have shown that there is a certain correlation between the activity (content) of markers in biological tissues and the content of pollutants in the environment. Therefore, studying the changes in the content of pollutants in the sediments of the East China Sea can provi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12N15/53C12N15/10C12Q1/6876
CPCC12N9/0065C12Q1/6876C12Q2600/158C12Y111/01006
Inventor 祁鹏志董文强任世太郭宝英
Owner ZHEJIANG OCEAN UNIV
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