Fluorescent quantitation PCR detection of Ehrlichia bacterium and typing primer, probe and kit

A technology for fluorescence quantification and detection of primers, which is applied in the determination/inspection of microorganisms, biochemical equipment and methods, DNA/RNA fragments, etc., to achieve good economic benefits, high specificity and sensitivity, and obvious technical advantages.

Inactive Publication Date: 2014-02-12
YANGZHOU UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0004] In order to solve the deficiency that the existing molecular diagnostic technology can only detect a single type of Ehrlichia, the present invention provides a rapid, specific, high-sensitivity quantitative PCR method suitable for clinical use to detect all kinds of Ehrlichia Primers, Probes and Kits

Method used

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  • Fluorescent quantitation PCR detection of Ehrlichia bacterium and typing primer, probe and kit
  • Fluorescent quantitation PCR detection of Ehrlichia bacterium and typing primer, probe and kit
  • Fluorescent quantitation PCR detection of Ehrlichia bacterium and typing primer, probe and kit

Examples

Experimental program
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Effect test

Embodiment 1

[0040] Example 1: Molecular identification of Ehrlichia

[0041] Molecular diagnosis was performed in 10 dogs with Ehrlichia who were clinically diagnosed, microscopically and serologically confirmed on blood smears. The operating procedure was as described above, and 400 μl of EDTA whole blood was used to extract DNA for PCR amplification. All 10 samples were PCR positive for Ehrlichia with the characteristic 63-64°C melting temperature (as Figure 5 shown). Gene sequencing confirmed the PCR results.

Embodiment 2

[0042] Example 2: Molecular epidemiological investigation of E. canis infection

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PUM

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Abstract

The invention discloses a fluorescent quantitation PCR detection of Ehrlichia bacterium and a typing primer, a probe and a kit. The sequence of the primer is shown in SEQ ID No. 1 and 2; the probe is shown in SEQ ID No. 3 and 4. The primer and the probe are adopted to carry out the PCR amplification for the bacterium to be detected and amplify 210bp band, and the fluorescent detection enhanced in 640 nanometers wavelength is positive samples of Ehrlichia bacterium; and then the Ehrlichia bacterium are typed according to the melting temperature. The invention has obvious technical advantages, high specificity and high sensitiveness, can conveniently detect all kinds of Ehrlichia bacteria, can be used for diagnosing and treating human and various animals (cattle, dogs and sheep) and Ehrlichia chafeensis diseases, and has good economic benefits.

Description

technical field [0001] The present invention relates to primers, probes and kits for fluorescent quantitative PCR detection and typing of molecular diagnosis of Ehrlichiella infection. Background technique [0002] Ehrlichia is an obligate intracellular parasite that lives in the cytoplasm of monocytes and neutrophils. Ehrlichiosis is transmitted by ticks and is widespread worldwide, causing a disease of humans and various animals (cattle, dogs, sheep) characterized by fever and anemia. The clinical diagnosis of the disease mainly relies on epidemiology and symptoms, combined with the microscopic observation of blood smears to detect Ehrlichia. The sensitivity of clinical diagnosis is low, and the existing serological diagnostic methods cannot distinguish the recovered from the clinical infection after infection. [0003] Important species of Ehrlichia that infect humans and animals include: canis E. canis, human E. chafeensis and E. ewingii, bovine E. ruminatium, sheep E....

Claims

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Application Information

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Patent Type & Authority Patents(China)
IPC IPC(8): C12Q1/68C12Q1/10C12N15/11
Inventor 王成明王志强许传灵
Owner YANGZHOU UNIV
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