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Molecular detection kit for detecting resistance of pig Escherichia coli F18 and application thereof

A technology of Escherichia coli and kits, applied in the field of molecular biology, can solve problems such as complicated operation and difficult sampling, and achieve the effect of strong repeatability, high accuracy and easy operation

Inactive Publication Date: 2010-06-09
YANGZHOU UNIV
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AI Technical Summary

Problems solved by technology

[0004] In view of the great harm caused by F18 E. coli in pig production, people have been looking for an accurate and rapid detection method for the identification of F18 E. coli resistance in pig production practice. The existing methods are mainly based on E. coli serotypes For the detection of virulence factors, sampling is difficult, the operation is relatively complicated, and there is a certain degree of subjectivity, so the application in breeding practice, epidemiological research and diagnosis has great limitations.

Method used

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  • Molecular detection kit for detecting resistance of pig Escherichia coli F18 and application thereof
  • Molecular detection kit for detecting resistance of pig Escherichia coli F18 and application thereof
  • Molecular detection kit for detecting resistance of pig Escherichia coli F18 and application thereof

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Embodiment 1

[0038] 1) PCR amplification of genomic DNA FUT1 gene M307

[0039] About 1.0 g of each test individual ear-collected tissue block was put into a 1.5 mL Eppendoff tube in an ice box, and genomic DNA was extracted by the conventional phenol-chloroform method. Use this kit ①-④ and ⑥ to carry out PCR amplification of α(1,2) fucosyltransferase (FUT1) gene, and the final PCR reaction system is established as follows:

[0040]

[0041] Reaction conditions: After the sample was denatured at 94°C for 5 minutes, it was amplified according to the following procedure: 94°C for 40s, 60°C for 40s, 72°C for 45s, after 32 cycles, 72°C for 10 minutes, and stored at 4°C. The PCR product was electrophoresed in 1% agarose gel, and after the end, the UVP gel imaging system was used to analyze and detect the amplification result, and the size of the amplified fragment should be 161bp ( figure 1 ).

[0042] 2) Enzyme digestion analysis of the amplification product of FUT1 gene M307

[0043] Th...

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Abstract

The invention relates to a molecular kit and detection method for accurately and quickly detecting the resistance of pig Escherichia coli F18. By using 7 kinds of working solutions and the detection method which are provided by the invention, on the basis of analyzing the polymorphism of the alpha (1, 2) fucosyltransferase (FUT1) gene M307, the resistance of the pig Escherichia coli F18 can be simply, conveniently and accurately determined, thereby realizing the large-scale detection on samples. The invention also provides a positive control which can achieve the aim of identifying the resistance of the pig Escherichia coli F18 at high sensibility and specificity, has simple and convenient operation, high accuracy and strong repeatability, and has significance for resistance of diarrhea of weanling pigs, breeding for hydropsy resistance and research and diagnosis of epidemiology.

Description

technical field [0001] The invention relates to a rapid detection device and a detection method for pig F18 Escherichia coli resistance, belonging to the technical field of molecular biology. Background technique [0002] In swine diseases, diarrhea and edema in piglets around weaning are caused by enterotoxigenic Escherichia coli [0003] (Enterotoxigenic Escherichia coli, ETEC) is the most common acute and fatal infectious disease caused by F18. The morbidity and mortality of the disease are high, and the cure rate is very low. The pathogenicity of ETEC F18 directly depends on whether there is a receptor corresponding to F18 cohesin in the brush border of piglet intestinal mucosal epithelial cells, that is, E.coli F18 receptor (ETEC F18R). Therefore, the study of candidate genes of ETEC F18R is of great significance for marker-assisted selection of resistant individuals and disease-resistant breeding in the future. Vogeli et al. (1997) showed that the FUT1 gene is the ET...

Claims

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Application Information

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IPC IPC(8): C12Q1/68
Inventor 包文斌吴圣龙叶兰黄小国朱国强潘章源朱璟孙寿永鞠慧萍艾晓峰何小亮
Owner YANGZHOU UNIV
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