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Multiplex PCR primer-probe group and kit for detecting pathogenic mucor

A technology of primer probe and primer set, which is applied in the direction of recombinant DNA technology, biochemical equipment and methods, microbial measurement/testing, etc., can solve the problems of poor specificity, interference, etc., and achieve high specificity, rapid detection, and detection sensitivity high effect

Pending Publication Date: 2022-01-07
HANGZHOU DIANZI UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

Using this kit, although one-time detection of multiple mucormycetes can be realized, since the primers are designed for the conserved region of the common gene of multiple mucormycetes, the specificity is poor, and it is easy to be affected by non-targets during the detection process. Bacteria (especially non-pathogenic mucormycetes of the same genus)

Method used

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  • Multiplex PCR primer-probe group and kit for detecting pathogenic mucor
  • Multiplex PCR primer-probe group and kit for detecting pathogenic mucor
  • Multiplex PCR primer-probe group and kit for detecting pathogenic mucor

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0036] Embodiment 1: kit and method of use

[0037]

[0038] A multiplex PCR kit for detecting pathogenic Mucormyces, comprising: PCR reaction buffer, magnesium chloride, deoxyribonucleoside triphosphate, Taq enzyme, uracil-N-glycosylase and primer probe set.

[0039] The primer probe set includes the following 3 pairs of upstream and downstream primers and 3 probes:

[0040] 1) Upstream primers, downstream primers and probes for detecting Rhizopus oryzae, the sequences of which are shown in SEQ ID NO.1, SEQ ID NO.2 and SEQ ID NO.7 respectively; the probes are labeled with FAM;

[0041] 2) Upstream primers, downstream primers and probes for detecting Rhizomucor micromyces, the sequences of which are shown in SEQ ID NO.3, SEQ ID NO.4 and SEQ ID NO.8 respectively; the probes are labeled with ROX;

[0042] 3) Upstream primers, downstream primers and probes for detecting A. umbelliferae, the sequences of which are shown in SEQ ID NO.5, SEQ ID NO.6 and SEQ ID NO.9 respectively;...

Embodiment 2

[0053] Embodiment 2: kit and method of use

[0054] A multiplex PCR kit for detecting pathogenic Mucormyces, including: PCR reaction buffer, magnesium chloride, deoxyribonucleoside triphosphate, Taq enzyme, uracil-N-glycosylase, primer probe set, negative Control substance, positive control substance and internal reference.

[0055] The primer probe set includes the following 4 pairs of upstream and downstream primers and 4 probes:

[0056] 1) Upstream primers, downstream primers and probes for detecting Rhizopus oryzae, the sequences of which are shown in SEQ ID NO.1, SEQ ID NO.2 and SEQ ID NO.7 respectively; the probes are labeled with FAM;

[0057] 2) Upstream primers, downstream primers and probes for detecting Rhizomucor micromyces, the sequences of which are shown in SEQ ID NO.3, SEQ ID NO.4 and SEQ ID NO.8 respectively; the probes are labeled with ROX;

[0058] 3) Upstream primers, downstream primers and probes for detecting A. umbelliferii, the sequences of which are...

Embodiment 3

[0068] Embodiment 3: kit and method of use

[0069] The difference between this example and Example 2 lies in: in step (2), the PCR reaction system was prepared according to Table 4; in step (3), the PCR reaction program shown in Table 5 was adopted.

[0070] Table 4 PCR reaction system

[0071]

[0072] Table 5 PCR reaction program

[0073]

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Abstract

The invention relates to the technical field of mucor detection and discloses a multiplex PCR primer-probe group and kit for detecting pathogenic mucor. The primer-probe group comprises a primer group and a probe group, wherein the primer group comprises an upstream primer and a downstream primer for detecting rhizopus oryzae, and sequences of the upstream primer and the downstream primer are separately shown as SEQ ID NO.1 and SEQ ID NO.2; an upstream primer and a downstream primer for detecting rhizomucor pusillus, wherein sequences of the upstream primer and the downstream primer are separately shown as SEQ ID NO.3 and SEQ ID NO.4; and an upstream primer and a downstream primer for detecting Lichtheimia corymbifera, wherein sequences of the upstream primer and the downstream primer are separately shown as SEQ ID NO.5 and SEQ ID NO.6. The primer-probe group and the kit, provided by the invention, can be used for achieving synchronous detection on the rhizopus oryzae, the rhizomucor pusillus and the Lichtheimia corymbifera at a species level, and have the advantages of high detection speed, high detection specificity and high detection accuracy.

Description

technical field [0001] The invention relates to the technical field of mucormycetes detection, in particular to a multiplex PCR primer probe set and kit for detecting pathogenic mucormycetes. Background technique [0002] Mucormycosis is an emerging serious infectious disease caused by the filamentous fungus Mucormyces, which develops rapidly and has a high mortality rate, mainly affecting immunocompromised patients and patients with diabetes. In fungal infections, the incidence of mucormyces accounts for 8.3% to 13%, second only to candida and aspergillus. In recent years, the incidence of mucormycosis in high-risk groups has increased, and it is about 1% in patients with malignant tumors and organ transplants. Mucormycosis is the second most common deep fungal infection in immunocompromised patients. According to reports, the incidence of mucormycosis in the United States and France increased by about 7% per year between 2000 and 2010, while the mortality rate increased ...

Claims

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Application Information

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IPC IPC(8): C12Q1/686C12Q1/6895C12Q1/04C12N15/11
CPCC12Q1/686C12Q1/6895C12Q2600/16C12Q2600/166C12Q2537/143C12Q2545/101C12Q2545/113C12Q2563/107
Inventor 应南娇莫秋思刘文佳
Owner HANGZHOU DIANZI UNIV
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