Looking for breakthrough ideas for innovation challenges? Try Patsnap Eureka!

Enterovirus type 71 (EV71) real-time fluorescent nucleic acid isothermal amplification detection kit

An enterovirus, EV71 technology, applied in the determination/inspection of microorganisms, biochemical equipment and methods, microorganisms, etc., can solve the problems of long detection cycle, false positive experimental results, and low sensitivity.

Active Publication Date: 2013-11-13
SHANGHAI RENDU BIOTECH
View PDF4 Cites 6 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0006] In order to solve the problems that the existing enterovirus 71 (EV71) detection method has low sensitivity, long detection cycle, easy to cause contamination of amplified products, false positive or false negative of experimental results and high detection cost, the present invention provides A real-time fluorescent nucleic acid constant temperature amplification detection technology for enterovirus 71 (EV71) with short detection cycle, high sensitivity, high specificity, low pollution, stable reaction and low detection cost, easy to popularize and apply, including special primers, probes Needles, kits and their use

Method used

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
View more

Image

Smart Image Click on the blue labels to locate them in the text.
Viewing Examples
Smart Image
  • Enterovirus type 71 (EV71) real-time fluorescent nucleic acid isothermal amplification detection kit
  • Enterovirus type 71 (EV71) real-time fluorescent nucleic acid isothermal amplification detection kit
  • Enterovirus type 71 (EV71) real-time fluorescent nucleic acid isothermal amplification detection kit

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0098] Embodiment 1, be used for the design of special primer and probe of real-time fluorescent nucleic acid constant temperature amplification detection enterovirus 71 type (EV71)

[0099] The present invention selects no secondary structure and a highly conserved segment in the EV71 virus VP1 gene as the amplified target sequence region (its nucleotide sequence is shown in sequence 1 in the sequence table), and according to the principle of primer probe design, DNAStar, DNAMAN Software and manual design are used for real-time fluorescent nucleic acid constant temperature amplification to detect the special primer and probe sequence of enterovirus 71 (EV71), obtain the following specific sequence:

[0100] (1) A capture probe (TCO, Target Capture O1igo) that can specifically bind to the target nucleic acid (EV71 RNA) sequence of enterovirus type 71 (EV71), the nucleotide sequence of the capture probe is: 5' AACCUGUUAUAUAUCUAUGUCCCAGUUGAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA3' (seque...

Embodiment 2

[0104] Embodiment 2, prepare the real-time fluorescent nucleic acid constant temperature amplification detection kit of enterovirus 71 (EV71)

[0105] Using the special primers and probes provided in Example 1, a real-time fluorescent nucleic acid constant temperature amplification detection kit for enterovirus 71 (EV71) of the present invention was obtained. The kit contains capture probe (TCO, Target Capture Oligo), T7 primer, nT7 primer, EV71 detection probe, internal standard detection probe, internal standard, M-MLV reverse transcriptase and T7 RNA polymerase etc. point.

[0106] The capture probe exists in the nucleic acid extraction solution, the T7 primer, nT7 primer, EV71 detection probe, and internal standard detection probe exist in the EV71 detection solution, and the M-MLV reverse transcriptase and T7 RNA polymerase The enzyme exists in the SAT enzyme solution. Specifically, the kit is divided into A box (specimen processing unit) stored at 2-30°C and B box (nucl...

Embodiment 3

[0139] Example 3. Real-time fluorescent nucleic acid constant temperature amplification detection of clinical sample throat swab

[0140] Use the kit of the present invention (see Example 2 for composition) to detect enterovirus 71 (EV71) in the throat swab of clinical samples, and the specific method comprises the following steps:

[0141] 1. Sample collection, transportation and storage

[0142] The clinician will collect the specimens according to the actual situation. The test specimens are throat swabs. The collection method is: wipe the posterior pharyngeal wall and the pharyngeal tonsils on both sides with a special sampling cotton swab, invade the swab into 3-5mL normal saline, and send it sealed for inspection. Within 48 hours after the sample is collected, store it at 4°C and send it to the enterovirus monitoring network laboratory (or store it at -70°C for testing, and deliver it within one week).

[0143] 2. Nucleic acid extraction

[0144] 2.1 Add 200 μl lysate ...

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

PUM

No PUM Login to View More

Abstract

The invention discloses an enterovirus type 71 (EV71) real-time fluorescent nucleic acid isothermal amplification detection kit comprising reagents such as a capturing probe, EV71 amplification primers T7 primer and nT7 primer, a EV71 detection probe, M-MLV reverse transcriptase, T7 RNA polymerase, and the like. The kit can be used for detecting EV71 RNA in throat swab or stool, and has the characteristics of high specificity, high sensitivity (reaching 10copies / reaction), low pollution (amplification product RNA can be easily degraded under natural environment), and fast detection (conventionally detection can be finished within 60min). The kit can perform important effect in clinical diagnosis of EV71 early-stage infection, and can be widely applied.

Description

technical field [0001] The invention relates to the technical field of biological detection of viruses, in particular to primers used in the real-time fluorescent nucleic acid constant temperature amplification detection of enterovirus type 71 (EV71), which combines specific target capture technology and real-time fluorescent nucleic acid constant temperature amplification detection technology, Probes and related kits. Background technique [0002] Hand, foot and mouth disease is a global infectious disease that mostly occurs in infants and young children. It can cause herpes on the hands, feet, and mouth. In a small number of patients, it can cause fatal complications such as myocarditis, pulmonary edema, and meningoencephalitis. In my country, hand, foot and mouth disease was first discovered in Shanghai in 1981; since then, it has been reported all over the country. In some areas the prevalence is as high as 1000 / 100,000. [0003] At present, there are more than 20 kind...

Claims

the structure of the environmentally friendly knitted fabric provided by the present invention; figure 2 Flow chart of the yarn wrapping machine for environmentally friendly knitted fabrics and storage devices; image 3 Is the parameter map of the yarn covering machine
Login to View More

Application Information

Patent Timeline
no application Login to View More
IPC IPC(8): C12Q1/70C12Q1/68C12N15/11C12R1/93
Inventor 于明辉马道亮居金良
Owner SHANGHAI RENDU BIOTECH
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Patsnap Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Patsnap Eureka Blog
Learn More
PatSnap group products

Browse by: Latest US Patents, China's latest patents, Technical Efficacy Thesaurus, Application Domain, Technology Topic, Popular Technical Reports.

© 2024 PatSnap. All rights reserved.Legal|Privacy policy|Modern Slavery Act Transparency Statement|Sitemap|About US| Contact US: help@patsnap.com