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Application of ND5-CD in traceability of poultry manure pollution

An ND5-CD, poultry manure technology, applied in the application field of poultry manure pollution traceability, can solve the problems of manure pollution source, affecting the timely management and control of water pollution source, etc.

Pending Publication Date: 2020-12-22
UNIVERSITY OF CHINESE ACADEMY OF SCIENCES
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Otherwise, it will cause misjudgment of the source of fecal pollution and affect the timely control of water pollution sources

Method used

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  • Application of ND5-CD in traceability of poultry manure pollution
  • Application of ND5-CD in traceability of poultry manure pollution
  • Application of ND5-CD in traceability of poultry manure pollution

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0022] collection of samples

[0023] In order to make the samples representative, a total of 148 samples were collected from 5 regions including Inner Mongolia, Beijing, Henan, Jiangxi and Guangdong, including 105 poultry feces samples and 43 non-poultry feces samples. Among them, 105 poultry samples included 75 chicken manure samples, 10 duck manure samples, 14 goose manure samples and 6 pigeon manure samples; 43 non-poultry manure samples included 10 pig manure samples, 6 dog manure samples, 6 horse feces samples, 5 donkey feces samples, 6 cow feces samples and 10 sheep feces samples. After the sample collection was completed, the samples were transported back to the laboratory as soon as possible using dry ice and stored in a -80°C refrigerator.

Embodiment 2

[0025] DNA extraction

[0026] ①Take 180-220 mg of fecal samples, and use the QIAamp DNA Stool Mini Kit (QIAGEN, Hilden, Germany) DNA extraction kit to extract DNA from the samples according to the instructions.

[0027] ② Use the kit OneStep TM The PCR Inhibitor Removal Kit (Zymo Research Corps, Irvine, CA, USA) removed inhibitors from the resulting DNA.

[0028] ③Use NaNodrop 2000 spectrophotometer (Thermo Fisher Technologies, Foster City, CA, USA) to measure the concentration of the obtained DNA.

Embodiment 3

[0030] An application of using ND5-CD for traceability of poultry manure pollution, detected by fluorescent quantitative PCR method, the detection primer (ND5-CD) is composed of ND5-F, ND5-R and ND5-P, and the primer sequence is:

[0031] ND5-F: 5'-ACCTCCCCCAACTAGC-3', shown in SEQ ID NO:1;

[0032] ND5-R: 5'-TTGCCAATGGTTAGGCAGGAG-3', shown in SEQ ID NO:2;

[0033] ND5-P: 5'-TCAACCCATGCCTTCTT-3', shown in SEQ ID NO:3.

[0034] Fluorescent quantitative PCR reaction system: 10 μL 2×TaqMan Environmental Master Mix 2.0, upstream primer ND5-F with a final concentration of 900 nM, downstream primer ND5-R with a final concentration of 900 nM, probe ND5-P with a final concentration of 250 nM, 2 μL DNA template, add sterile water to 20μL.

[0035] Fluorescence quantitative PCR reaction conditions: pre-denaturation at 50°C for 2 min; pre-denaturation at 95°C for 10 min; denaturation at 95°C for 15 s, annealing and extension at 60°C for 1 min, a total of 40 cycles.

[0036] Poultry in...

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Abstract

The invention discloses an application of ND5-CD in traceability of poultry manure pollution, and relates to the technical field of microorganism traceability. The marker ND5-CD is originally only used for manure pollution detection of chickens and ducks, and in the invention, the marker ND5-CD is innovatively applied to poultry manure pollution detection on the basis of in-depth assessment of applicable microorganism traceability objects of the marker, so that source of manure pollution is accurately assessed.

Description

technical field [0001] The invention relates to the technical field of microbial traceability, in particular to an application of ND5-CD for traceability of poultry manure pollution. Background technique [0002] With the improvement of people's living standards, the scale of poultry farming industry is constantly expanding. According to the statistics of "China Animal Husbandry and Veterinary Yearbook 2016", the number of poultry farming ranks first in my country. The large amount of feces brought by poultry farming is likely to cause water pollution incidents. At the same time, poultry feces contain a large number of pathogenic bacteria, and contact with untreated fecal water can easily cause human illness and pose a potential threat to human health. Therefore, rapid and accurate identification of feces from poultry is of great significance to water protection and human health. [0003] Microbial traceability is a technology that uses the characteristics of specific seq...

Claims

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Application Information

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IPC IPC(8): C12Q1/689C12Q1/6895C12Q1/686C12N15/11
CPCC12Q1/689C12Q1/6895C12Q1/686
Inventor 余志晟梁红霞张洪勋
Owner UNIVERSITY OF CHINESE ACADEMY OF SCIENCES
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