Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Molecular marker capable of rapidly detecting drug resistance of chicken coccidiosis, detection method and kit

A technology of molecular markers and drug resistance, applied in the fields of botany equipment and methods, biochemical equipment and methods, microbial measurement/inspection, etc., can solve the problems of long test cycle, long detection cycle, high cost, etc.

Pending Publication Date: 2021-11-02
SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
View PDF0 Cites 0 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

This method is cumbersome, time-consuming and laborious, and requires well-trained experts to make judgments. Due to the long test period, it will largely lead to delays in the treatment of coccidiosis on-site
[0005] In short, the above-mentioned commonly used diagnostic methods for coccidia drug resistance have problems such as long detection period, high cost, cumbersome operation, and the need for well-trained experts; the market needs a rapid, specific, and highly sensitive detection method. The present invention solves such problems

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Molecular marker capable of rapidly detecting drug resistance of chicken coccidiosis, detection method and kit
  • Molecular marker capable of rapidly detecting drug resistance of chicken coccidiosis, detection method and kit
  • Molecular marker capable of rapidly detecting drug resistance of chicken coccidiosis, detection method and kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0053] Example 1 Screening and biological analysis of coccidia drug resistance molecular markers

[0054] The three-yellow chickens needed for this study were all purchased from a poultry farm in the suburbs of Fengxian, Shanghai, and the 1-day-old chicks were raised in a strict coccidia-free environment.

[0055] The sensitive strain of Eimeria tenella used in this experiment was isolated from chickens on a farm in Shanghai in the 1980s, and was bred and preserved by the laboratory (resource number: CAAS21111601).

[0056] In the early stage, the laboratory adopted the method of gradually increasing the drug concentration, with 0.05ppm diclazuril and 2.0ppm maduramycin as the initial induction concentration, and after 18 and 20 passages respectively, the tender Eimeria spp. was induced in the laboratory Resistant strains of worms to diclazuril and maduramycin. Wherein the diclazuril-resistant strain has complete resistance to 1.2ppm diclazuril, and is fully sensitive to madu...

Embodiment 2

[0068] Example 2 Detection of coccidia drug resistance molecular markers

[0069] First, by subcutaneous injection, the purified r Et ENO2, r Et PGK and r AKRO immunizes mice and rabbits to obtain polyclonal antibodies against mouse and rabbit anti-recombinant proteins. For specific steps, please refer to reference books such as "Molecular Cloning".

[0070] Second, the drug resistance-associated genes were determined by relative fluorescence quantitative PCR Et ENO2 、Et PGK and Et The mRNA transcription level of AKRO at different developmental stages of the life cycle of sensitive strains, with 18s rRNA as the control. For specific operations, please refer to reference books such as Molecular Cloning. Summarized as:

[0071] Preparation of the template:

[0072] 1) Extraction of total RNA. The RNA of four different developmental stages (unsporulated oocysts, sporulated oocysts, sporozoites, and second-generation merozoites) of Eimeria tenena sensitive strains was e...

Embodiment 3

[0113] Example 3 Application of coccidia drug resistance molecular markers in the process of preventing parasites from invading cells

[0114] Referring to "Molecular Cloning" and other reference books, the invasion experiment was carried out to test the effect of the rabbit anti-recombinant protein polyclonal antibody on the invasion of sporozoites into DF-1 cells.

[0115] 1) Take out the frozen DF-1 cells and place them in a 37°C water bath to dissolve them quickly, then suck them into a 15mL centrifuge tube, and collect the precipitate by centrifugation. Cells were resuspended in cell culture medium containing 10% fetal bovine serum, then transferred to cell culture flasks and cultured in a cell culture incubator.

[0116] 2) Subculture when the cell density is high. Pour off the original culture medium in the culture bottle, wash it twice with PBS, and then digest the adherent cells with 1~2mL trypsin, and immediately add 10mL of cell culture medium containing 10% fetal ...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

PropertyMeasurementUnit
Isoelectric pointaaaaaaaaaa
Isoelectric pointaaaaaaaaaa
Login to View More

Abstract

The invention relates to a molecular marker capable of rapidly detecting drug resistance of chicken coccidiosis, a detection method and a kit. In the invention, three drug resistance related genes, namely EtENO2, EtPGK and EtAKRO, are cloned. The mRNA transcriptional level difference and the protein translation level difference of two drug-resistant strains (diclazuril drug-resistant strains and maduramycin drug-resistant strains) and drug-sensitive strains induced by three genes in a laboratory are compared, and meanwhile, the drug resistance of field isolated strains is detected, and a theoretical basis is provided for further researching a molecular mechanism of coccidiosis drug resistance generation and establishing rapid detection of field coccidiosis drug resistance in the future.

Description

technical field [0001] The invention relates to the field of disease detection assistance, in particular to a molecular marker, a detection method and a kit for rapidly detecting chicken coccidia drug resistance. Background technique [0002] Chicken coccidiosis is a very common fatal epidemic caused by several kinds of Eimeria coccidiosis parasitic chicken intestines, which seriously endangers the growth and development of chickens, and is one of the most harmful diseases in intensive chicken farming. The disease causes huge economic losses to the global poultry industry every year. Due to the poor stability of coccidiosis vaccines, the current prevention and control of coccidiosis mainly relies on drugs. With the long-term use of anticoccidiosis drugs in large-scale chicken farms, coccidia are prone to drug resistance, and chicken farm balls The problem of insecticide resistance has become very serious. Since 1954, after the United States isolated strains resistant to su...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12Q1/6893C12Q1/686C12Q1/6806C12N15/60C12N15/53C12N15/54
CPCC12Q1/6893C12Q1/686C12Q1/6806C12N9/88C12N9/1217C12N9/0004C12Y207/02003C12Q2600/106C12Q2600/156C12Q2563/107C12Q2545/114
Inventor 韩红玉董辉黄文浩黄兵赵其平朱顺海于钰姚亚文刘展
Owner SHANGHAI VETERINARY RES INST CHINESE ACAD OF AGRI SCI
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com