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Detection kit for HPIV (human parainfluenza virus) triple nucleic acid

A technology of human parainfluenza virus and detection kit, which is applied in the field of nucleic acid detection, can solve the problems of poor specificity and low sensitivity of the kit, and achieve the effect of fast operation, high sensitivity and low resolution efficiency

Inactive Publication Date: 2018-12-28
AUTOBIO DIAGNOSTICS CO LTD
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  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The object of the present invention is to provide a human parainfluenza virus triple nucleic acid detection kit to solve the technical problems of low sensitivity and poor specificity of the kit in the prior art

Method used

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  • Detection kit for HPIV (human parainfluenza virus) triple nucleic acid
  • Detection kit for HPIV (human parainfluenza virus) triple nucleic acid
  • Detection kit for HPIV (human parainfluenza virus) triple nucleic acid

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0017] Example 1 Preparation of Human Parainfluenza Virus Triple Nucleic Acid Detection Kit

[0018] Prepare the PCR reaction system according to the ratio in Table 2

[0019] Table 2

[0020]

[0021] This kit also includes a negative control (sterile water) and a positive control (artificially synthesized at a concentration of 1×10 5 Copies / ml of pseudovirus).

Embodiment 2

[0022] Embodiment 2 The detection method of kit of the present invention

[0023] The detection method of the present invention is Real time RT-PCR, which combines RNA reverse transcription and DNA amplification. The Real TimeRT-PCR reaction process is (1) cDNA synthesis; (2) pre-denaturation, the time and length depend on the length and base composition of the target nucleotide, the pre-denaturation temperature is generally 90°C-105°C, and the time is generally 1 -10min, the purpose of pre-denaturation is to completely separate the double-stranded nucleotide sequence into single strands; (3) Denaturation, the temperature is generally 90°C-105°C, and the time is generally 10s-30s; (4) Annealing, so that each primer Anneal to the target sequence of human parainfluenza virus type 1, human parainfluenza virus type 2, human parainfluenza virus type 3 or internal standard quality control nucleic acid. The annealing temperature is usually 40°C-60°C, and the annealing time can be 10...

Embodiment 3

[0033] Embodiment 3 Feasibility test of kit of the present invention

[0034] 1. Lower limit of detection (LOD) test

[0035] (1) Preparation of human parainfluenza virus triple nucleic acid detection reagent

[0036] The human parainfluenza virus triple nucleic acid detection reagent was prepared by adopting the method of Example 1.

[0037] (2) Virus sample extraction

[0038] Mix the pharyngeal swab eluate of five different concentrations of human parainfluenza virus type 1, type 2, and type 3 in the tube with a pipette, take out 200 µl into a new centrifuge tube, centrifuge at 12,000 rpm for 5 min, and discard carefully Remove the supernatant; add 200 µl of virus lysate to the pellet, mix well, bathe in water at 100°C for 5 min, and centrifuge at 10,000 rpm for 5 min for later use.

[0039] (3) Sample detection

[0040] Add 25µl of the processed specimen supernatant to the human parainfluenza virus triple nucleic acid detection reaction tube, 20 duplicate wells for eac...

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Abstract

The invention discloses a detection kit for HPIV (human parainfluenza virus) triple nucleic acid. The detection kit is characterized by comprising HPIV type 1, type 2 and type 3 nucleic acid PCR reaction liquid, wherein the PCR reaction liquid comprises reaction buffer liquid, deoxyribonucleoside triphosphate, upstream and downstream primers for amplification of HPIV type 1, type 2 and type 3 target polynucleotides and probes for detection of target polynucleotides, and the sequence table of the probes is shown as SEQ ID NO:1-9. The detection kit has the advantages of being rapid, convenient and simple to operate, good in detection specificity, high in sensitivity and wide in detection range. As the probes with higher specificity is applied to the kit, the kit can rapidly detect the HPIV type 1, type 2 and type 3 nucleic acids in unknown samples; reliable experimental basis is provided for diagnosing the HPIV type 1, type 2 and type 3 nucleic acids, and the technical problems of low efficiency, poor specificity and low sensitivity of existing kits are solved.

Description

technical field [0001] The invention relates to nucleic acid detection technology, in particular to a human parainfluenza virus triple nucleic acid detection kit. Background technique [0002] Human parainfluenza virus (HPIV), formerly known as Sendai virus, is a circular closed, antisense RNA virus belonging to the family Paramyxoviridae. According to genetic and antigenicity can be divided into 4 types (HPIV1-4). Among them, HPIV-1 and HPIV-3 are often the main causes of respiratory infection outbreaks in public places. HPIV-3 is second only to respiratory syncytial virus (RSV), and it can especially cause acute respiratory infection (ARI) in infants and young children. , HPIV-1 and 2 mainly cause laryngotracheobronchial inflammation. HPIV-4 is further divided into two subtypes, A and B, according to different antigenicities. HPIV4 is generally considered to exist only sporadically, and the infection symptoms caused are generally mild, and rarely cause outbreaks of respi...

Claims

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Application Information

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Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/6851C12R1/93
CPCC12Q1/6851C12Q1/701C12Q2600/16C12Q2600/166C12Q2531/113C12Q2537/143C12Q2545/101C12Q2521/531
Inventor 高歌高利飞杜美李振红付光宇吴学炜苗拥军
Owner AUTOBIO DIAGNOSTICS CO LTD
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