MNP marker site of mumps virus, primer composition, kit and application of MNP marker site

A technology of mumps and primer combination, applied in the biological field, can solve the problems of small number, huge number of SNP markers, insufficient to capture allelic diversity, etc., and achieve the effect of high-accuracy and high-sensitivity detection

Active Publication Date: 2022-08-02
JIANGHAN UNIVERSITY
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

SSR markers are recognized as the most polymorphic markers, but their number is small in microorganisms; the number of SNP markers is huge, densely distributed, and they are dimorphic markers, and the polymorphism of a single SNP marker is not enough to capture potential alleles in microbial populations genetic diversity

Method used

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  • MNP marker site of mumps virus, primer composition, kit and application of MNP marker site
  • MNP marker site of mumps virus, primer composition, kit and application of MNP marker site
  • MNP marker site of mumps virus, primer composition, kit and application of MNP marker site

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0044] Example 1. Screening of mumps virus MNP marker sites and design of multiple PCR amplification primers

[0045] S1. Screening of mumps virus MNP marker sites

[0046] Based on the complete or partial genome sequences of 3794 different isolates of mumps virus published on the Internet, 15 MNP marker sites were obtained through sequence alignment. For species for which there is no genome data online, the genome sequence information of the representative race of the microbial species to be detected can also be obtained by high-throughput sequencing, where the high-throughput sequencing can be whole genome or simplified genome sequencing. In order to ensure the polymorphism of the selected marker, the genome sequences of at least 10 genetically representative isolates are generally used as a reference. The 15 MNP marker sites screened are shown in Table 1:

[0047] Table 1 - The MNP marker site and the starting position of the detection primer on the reference sequence

...

Embodiment 2

[0060] Threshold setting and performance evaluation of MNP markers and primers for identification of mumps virus described in Example 2

[0061] 1. Detection of MNP markers

[0062] Use the purchased mumps virus RNA quality control material with known copy number (goods number: VIP(VC)040), after reverse transcribing into cDNA by a commercial reverse transcription kit, add it to human genomic DNA, Mumps virus mock samples were prepared at 1 copy / reaction, 10 copies / reaction and 100 copies / reaction. At the same time, an equal volume of sterile water was set as a blank control. A total of 4 samples were collected, and 3 replicate libraries were constructed for each sample every day, and were continuously detected for 4 days, that is, 12 sets of sequencing data were obtained for each sample, as shown in Table 2. According to the number of mumps virus MNP-labeled sequencing fragments and loci detected in the blank control and the mumps virus nucleic acid quality control material...

Embodiment 3

[0085] Example 3. Detection of genetic variation among mumps virus strains

[0086] The kit and the MNP marker site detection method were used to detect 6 copies of the mumps virus strain in different periods provided by Hubei CDC. The samples were named S1-S6 in turn. The average coverage of the samples was 3641 times, and all 15 MNP markers could be detected for each strain (Table 5). The fingerprints of the 6 strains were compared in pairs, and the results were shown in Table 5. The other 5 mumps viruses detected by S3 and the same batch were all different in the main genotypes of 5 MNP markers ( Table 5), indicating that there is inter-strain variation.

[0087] Table 5-6 Detection and Analysis of Mumps Viruses

[0088]

[0089]

[0090] As can be seen from Table 5, the test kit of the present invention can be used to ensure the genetic consistency of the same named mumps virus strains in different laboratories by detecting the application of MNP markers to identif...

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Abstract

The invention discloses MNP marker sites of epidemic parotitis virus, a primer composition, a kit and application of the MNP marker sites, the primer composition and the kit, the MNP marker sites are genome areas which are screened from epidemic parotitis virus genome, are distinguished from other species and have multiple nucleotide polymorphisms in strains in the species, and comprise marker sites of MNP-1 to MNP-15; the primers are as shown in SEQ ID NO. 1 to SEQ ID NO. 30. The MNP marker site can be used for specifically identifying the mumps virus and distinguishing different strains; the primers do not interfere with one another, multiple amplification and sequencing technologies are integrated, and sequence analysis can be carried out on all marker sites of multiple samples at a time; the kit has the detection advantages of high throughput, multiple target points, high sensitivity, variation monitoring and culture-free detection, can be applied to detection of large-scale samples, and has important significance on scientific research and epidemic prevention monitoring of epidemic parotitis viruses.

Description

technical field [0001] The embodiments of the present invention relate to the field of biotechnology, in particular to an MNP marker site of mumps virus, a primer composition, a kit and applications thereof. Background technique [0002] Mumps virus (Mumps virus) is an RNA virus. Humans are the only host of mumps virus. The virus is spread by droplets. The susceptible people are school-age children. The main clinical manifestations are one or both parotid gland enlargement, accompanied by fever, fatigue, muscle pain, etc. About 0.1% of children can be complicated by viral meningitis. Mumps is also a common cause of acquired deafness in childhood. Vaccination is an effective preventive measure. After mumps disease, a firm immunity can be obtained. There is only one serotype of mumps virus, and the variation of the strain will affect the diagnostic scheme of clinical medicine. In addition, for the experimental study of mumps virus, the variation of the strain will lead to ...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/6858C12Q1/6869C12N15/11G16H50/80C12R1/93
CPCC12Q1/701C12Q1/6858C12Q1/6869G16H50/80C12Q2600/16C12Q2600/156C12Q2531/113C12Q2537/143C12Q2535/122Y02A50/30
Inventor 高利芬李甜甜李论陈利红周俊飞方治伟彭海肖华锋
Owner JIANGHAN UNIVERSITY
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