Multiple PCR (polymerase chain reaction) detection kit and method for giardia and cryptosporidia of cattle

A detection kit and cryptosporidium technology, applied in biochemical equipment and methods, microbiological measurement/inspection, resistance to vector-borne diseases, etc. High-level problems, to achieve the effect of convenient observation results, improved sensitivity and accuracy, and strong specificity

Inactive Publication Date: 2018-11-13
JILIN UNIV
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  • Application Information

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Problems solved by technology

[0004] At present, there are mainly three detection methods for Cryptosporidium amnesii, Cryptosporidium parvum and Giardia duodenum, which are pathogenic detection methods, immunological detection methods and molecular biological detection methods. The most widely used method is the pathogenic detection method, which d

Method used

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  • Multiple PCR (polymerase chain reaction) detection kit and method for giardia and cryptosporidia of cattle
  • Multiple PCR (polymerase chain reaction) detection kit and method for giardia and cryptosporidia of cattle
  • Multiple PCR (polymerase chain reaction) detection kit and method for giardia and cryptosporidia of cattle

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Embodiment 1

[0048] Specific gene selection of Giardia duodenum, Cryptosporidium parvum and Cryptosporidium ahnii

[0049] Species-specific diagnostic sequences of Giardia duodenum, Cryptosporidium parvum, and Cryptosporidium ahnii: select highly conserved TPI gene and 18S rRNA gene sequences, and compare them to Giardia and Cryptosporidium in NCBI It is unique to insects and meets the standards for species-specific detection and research.

[0050] Primer Design for Triple PCR Detection Kit

[0051] Use biological software to design specific diagnostic primers as follows:

[0052]Giardia duodenum PCR primers:

[0053] F1: 5'-CGACCTGACTATGACCAACTC-3';

[0054] R1: 5'-GTCCTGAAGCATTCTCAACACT-3';

[0055] Cryptosporidium parvum PCR primers:

[0056] F2: 5'-CCTTACTCCTTTCAGCACCTTATG-3';

[0057] R2: 5'-TGTTACGACTTCTCCTTCCTCTAA-3';

[0058] PCR primers for Cryptosporidium amnesia:

[0059] F3: 5'-TACCGTGGCTATGACGGGTA-3';

[0060] R3: 5'-CACCAGACTTGCCCTCCAAT-3';

[0061] A diagnostic gen...

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Abstract

The invention discloses a multiple PCR (polymerase chain reaction) detection kit and method for giardia and cryptosporidia of cattle. According to parts of conserved sequences of giardia duodenalis TPI genes, cryptosporidium parvum 18S rRNA genes and cryptosporidium andersoni 18S rRNA genes, a primer is designed, a triple PCR detection method is built, sensitivity and accuracy of the method are improved by optimizing reaction conditions such as annealing temperature of PCR reaction, amplification products are implemented by 1.0% of agarose gel electrophoresis, and a result is conveniently andrapidly observed. Three target fragments amplified by the method have size differences in agarose gel electrophoresis and are easily distinguished, the sensitivity can reach 1*10<3>/g and is higher than that of a reported microscopy method and a conventional PCR detection method of giardia duodenalis, cryptosporidium parvum and cryptosporidium andersoni, and the method is suitable for detection ofsamples in clinical practice. The detection method can simultaneously and rapidly detect the giardia duodenalis, the cryptosporidium parvum and the cryptosporidium andersoni of the cattle and has theadvantages of simplicity in operation, high sensitivity, high specificity and the like, and technical support is provided for quality standards of real experiment cattle.

Description

technical field [0001] The invention provides a method for simultaneously detecting bovine Giardia duodenum ( Giardia duodenalis, G. duodenalis ), Cryptosporidium parvum ( C. parvum) and Cryptosporidium angsleri ( C. Andersoni ) triple PCR detection method, the invention also provides the preparation method and usage of the kit, belonging to the technical field of parasite detection. Background technique [0002] Giardia ( Giardia ) and Cryptosporidium ( Cryptosporidium ) belongs to important zoonosis protozoa, Cryptosporidium includes Cryptosporidium parvum ( C. parvum) and Cryptosporidium amnesii ( C. Andersoni ), etc., mainly cause digestive tract and respiratory tract diseases in young animals, resulting in decreased production performance or death. Giardia is divided into 6 species, including Agile Giardia ( Giardia agi-lis ), Giardia heron ( Giardia ardeae ), Giardia parrot ( Giardia psittaci ), Giardia microns ( Giardia microti ), Giardia rat ( ...

Claims

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Application Information

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IPC IPC(8): C12Q1/6893C12Q1/686
CPCC12Q1/686C12Q1/6893C12Q2600/16C12Q2537/143Y02A50/30
Inventor 宫鹏涛张西臣高宇航李显赫任文陟李建华杨举
Owner JILIN UNIV
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