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Primer and probe for detecting Zika virus

A Zika virus and probe technology, applied in biochemical equipment and methods, microorganisms, and methods based on microorganisms, etc., can solve the problems of missed treatment time, difficult diagnosis of patients, long time, etc., to achieve optimal concentration, effective detection, And the effect of accurate detection

Pending Publication Date: 2021-04-02
SUN YAT SEN UNIV
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

It takes a long time to isolate and culture the virus, and there are serological cross-reactions among members of the Flaviviridae family. It is difficult to diagnose some patients infected with the virus within a short period of time, resulting in missing the best time for treatment.

Method used

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  • Primer and probe for detecting Zika virus
  • Primer and probe for detecting Zika virus
  • Primer and probe for detecting Zika virus

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0029] The design of embodiment 1 primer and probe

[0030] 1. Experimental method

[0031] By comparing and analyzing the known Zika virus gene sequences in all GenBank databases, selecting highly conserved segments, and verifying the specificity of the primers and the homology of similar pathogens by BLAST, a detection method for the above pathogens was designed. primers and probes.

[0032] 2. Experimental results

[0033] Primers and probes for Zika virus E gene were designed.

[0034] Wherein, the nucleotide sequence of the upstream primer is the nucleotide sequence shown in SEQ ID NO: 1,

[0035] The nucleotide sequence of the downstream primer is shown in SEQ ID NO: 2,

[0036] The nucleotide sequence of the specific probe is shown in SEQ ID NO: 3, its 5' end is fluorescently labeled with FAM, and its 3' end is fluorescently labeled with BHQ1.

Embodiment 2

[0037] Embodiment 2 A kind of kit for detecting Zika virus

[0038] 1. Composition

[0039] A primer whose nucleotide sequence is shown in SEQ ID NO: 1-2;

[0040] The nucleotide sequence is shown in SEQ ID NO: 3, the 5' end is fluorescently labeled with FAM, and the 3' end is fluorescently labeled with BHQ1 probe;

[0041] SEQ ID NO: 1: CCGCTGCCCAACACAAG,

[0042] SEQ ID NO: 2: CCACTAACGTTCTTTTGCAGACAT,

[0043] SEQ ID NO: 3: AGCCTACCTTGACAAGCAGTCAGACACTCAA.

[0044] 2. How to use

[0045] 1. Total RNA extraction

[0046] (1) Add 200ul RNAiso Plus or Triziol (the amount used when extracting RNA from a whole Aedes mosquito) to a 2ml RNase-free centrifuge tube, homogenize at 30Hz for 30s. Transfer the liquid to a 1.5ml RNase-free centrifuge tube and let stand for 15min.

[0047](2) Add 1 / 5 volume (40 μl) of chloroform to the above lysate, close the cap of the centrifuge tube tightly, and vortex to mix. After the solution was fully emulsified (no phase separation), let s...

Embodiment 3

[0071] The detection of embodiment 3 positive samples

[0072] 1. Experimental method

[0073] The kit of Example 2 was used to detect Aedes albopictus infected with Zika virus. At the same time, virus titer titration (TCID 50 / ml), TCID 50 That is half the infectious dose of tissue cell culture, with 1TCID 50 / ml of virus infects Aedes albopictus. One Aedes albopictus infected with Zika virus was used as the sample to be tested, and the kit in Example 2 was used for detection.

[0074] 2. Experimental results

[0075] See the test results figure 1 , only one amplification curve shows that the above primers and probes have good specificity.

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PUM

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Abstract

The invention provides a primer and a probe for detecting Zika virus. The primer and the probe are designed in a relatively conservative region, and the primer and the Taqman probe with high specificity are selected, so that the concentrations of the primer and the probe in a reaction system are optimized, the Zika virus in aedes can be accurately detected, and the Zika virus in the aedes can be rapidly, effectively and accurately detected; The primer and the probe can also be used for detecting clinical specimens.

Description

technical field [0001] The invention relates to the technical field of virus detection, in particular to a primer and a probe for detecting Zika virus. Background technique [0002] Zika virus (ZIKV) is a mosquito-borne virus belonging to the Flaviviridae family, and its main vectors are Aedes aegypti and Aedes albopictus. Aedes is an insect of the genus Aedes in the subfamily Culexidae of the mosquito family Culex, and mosquitoes in the genus Aedes are commonly known as Aedes mosquitoes. Aedes is a small to medium-sized black mosquito species with silvery white markings. There is a central white longitudinal stripe on the mid-thorax scutellum, which extends from the front end to the front of the small scutellum at the level of the wing base and forks. The 1st to 4th segments of the posterior tarsus have base white rings, and the distal segments are all white. There are basal leucorrhea in 2-6 segments on the back of the abdomen. The total length of the Zika virus genome...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/6851C12N15/11C12R1/93
CPCC12Q1/701C12Q1/6851C12Q2531/113C12Q2561/101
Inventor 黎孟枫李隽朱勋何振健吴珏珩谭姹辉陈德林林翠姬谭泳谣曹开源袁洁蔡俊超
Owner SUN YAT SEN UNIV
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