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Herpes simplex virus type I detection marker, primer probe pair, kit and detection method

A technology of virus detection and kit, applied in biochemical equipment and methods, measurement/inspection of microorganisms, DNA/RNA fragments, etc., can solve the problem of inability to accurately reflect whether infection or carrying pathogens, long time period for virus isolation and culture, HSV Problems such as difficulty in typing and detection, achieve low nucleic acid similarity, improve detection sensitivity, and not easily cross-react

Inactive Publication Date: 2019-03-29
中生方政生物技术股份有限公司
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  • Application Information

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Problems solved by technology

Virus isolation and culture takes a long period of time, cumbersome operations, professional operators and harsh laboratory conditions are required, so it is rarely used in clinical rapid diagnosis
[0008] 2) There is still lag in immunological diagnosis, which cannot accurately reflect whether the current infection or pathogen is carried, and it is difficult to meet the requirements of early diagnosis
The high similarity between HSV-1 and HSV-2 makes it difficult to detect HSV types

Method used

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  • Herpes simplex virus type I detection marker, primer probe pair, kit and detection method
  • Herpes simplex virus type I detection marker, primer probe pair, kit and detection method
  • Herpes simplex virus type I detection marker, primer probe pair, kit and detection method

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Embodiment 1

[0069] Embodiment 1: be used for the design of the primer probe pair of rapid detection HSV-1

[0070] For the HSV-1RL2 gene HBB gene sequence in NCBI, design primer pairs and probes, the sequence is as follows:

[0071] Table 1 Primer and Probe Sequences

[0072]

Embodiment 2

[0073] Embodiment 2: be used for the real-time fluorescent quantitative PCR kit of rapid detection HSV-1

[0074] Real-time fluorescent quantitative PCR kit for rapid detection of human HSV-1, including HSV-1 primers, HSV-1 probes, HBB primers, HBB probes, enzyme mixture, reaction buffer, sample extract, positive control , negative control substance, instructions and box body.

[0075] The sequences of primers HSV-1-F and HSV-1-R are as shown in SEQ ID NO.1-2 in sequence, and the concentration is 500nM; the sequence of probe HSV1-P is as shown in SEQ ID NO:3, and the fluorescent group is Cy5. The quenching group was BHQ3 at a concentration of 200 nM.

[0076] The sequences of primers HBB-F and HBB-R are shown in SEQ ID NO.4-5 in turn, and the concentration is 500nM; the sequence of probe HBB-P is shown in SEQ ID NO:6, and the fluorescent group of the probe is ROX, which is quenched The group is BHQ1 and the concentration is 200 nM.

[0077] Among them, Anstart qPCR Master M...

Embodiment 3

[0085] Embodiment 3: the rapid detection method of HSV-1 nucleic acid detection kit

[0086] Utilize the kit of embodiment 2 to quickly detect HSV-1 in human vaginal secretions, urinary tract secretions, blister fluid samples, and the specific steps are as follows:

[0087] (1) Nucleic acid extraction: 4 cases of vaginal secretions (sample numbers 1-4), 3 cases of urinary tract secretions (sample numbers 5-7), 3 cases of blister fluid samples (sample numbers 8-10) (using virus Separation culture and typing method to determine whether it is infected by HSV-1, this method is the current industry gold standard) Add 1mL of normal saline to the collection tube, fully shake and wash the cotton swab, then squeeze the cotton swab against the wall and discard it, take 500μL Transfer the liquid to a 1.5mL centrifuge tube, centrifuge at 13000rpm for 5 minutes, discard the supernatant, add 1mL of normal saline to the precipitate, break up the precipitate, centrifuge at 13000rpm for 5 minu...

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Abstract

The present invention relates to the technical field of detection of pathogenic microorganisms and particularly relates to a herpes simplex virus type I detection marker, a primer probe pair, a kit and a detection method. A reverse repeat sequence RL2 in a HSV genome is used as a target region,2 copies are in the genome, detection sensitivity can be further improved, similarity between nucleic acids of two types is low, and cross-reaction is not easy to cause.

Description

technical field [0001] The invention relates to the technical field of detection of pathogenic microorganisms, in particular to a herpes simplex virus type I detection marker, a primer-probe pair, a kit and a detection method. Background technique [0002] Herpes simplex is a viral skin disease caused by herpes simplex virus, which can cause various human diseases, such as gingivostomatitis, keratoconjunctivitis, encephalitis, reproductive system infection and neonatal child's infection. [0003] Herpes simplex virus (HSV) belongs to the subfamily Avirinae of the family Herpesviridae, and the size of the virus plasmid is about 180 nanometers. According to the difference in antigenicity, the virus is currently divided into type I (HSV-1) and type II (HSV-2). HSV-1 is mainly obtained from lip lesions, and HSV-2 can be isolated from genital lesions. Infection is due to human-to-human contact, and it is the easiest virus to infringe on human beings, but only a part of it deve...

Claims

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Application Information

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IPC IPC(8): C12Q1/70C12Q1/6851C12N15/11
CPCC12Q1/6851C12Q1/705C12Q2600/166C12Q2531/113C12Q2563/107C12Q2545/113C12Q2545/101
Inventor 邹国宝蔺皓魏颖颖刘欣欣宋高尚沈江卫吴茜
Owner 中生方政生物技术股份有限公司
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