Porcine circovivus loop-mediated isothermal amplification kit and application thereof

A porcine circovirus and ring-mediated isothermal technology, applied in the field of microbial detection, can solve the problems of expensive equipment, difficulty in detecting porcine circovirus, and long time consumption, and achieve the effect of simple operation, avoiding pollution, and rapid acquisition

Inactive Publication Date: 2015-05-27
GUANGXI VETERINARY RES INST
View PDF3 Cites 9 Cited by
  • Summary
  • Abstract
  • Description
  • Claims
  • Application Information

AI Technical Summary

Problems solved by technology

[0005] The purpose of the present invention is to solve the problems such as difficult, time-consuming and expensive instruments for detecting porcine circovirus at the grassroots level, and to provide a simple, fast and accurate test kit for detecting porcine circovirus at the grassroots level. The technical solution is: a porcine circovirus ring-mediated isothermal amplification kit, which includes LAMP primers, 2× reaction buffer, Bst DNA polymerase, fluorescent visual detection reagents, ultrapure water and porcine circovirus DNA template; the LAMP primers include outer primers F3 (SEQ ID NO: 1) and B3 (SEQ ID NO: 2), inner primers FIP (SEQ ID NO: 3) and BIP (SEQ ID NO: 4) and loop Primers LF (SEQ ID NO: 5) and LB (SEQ ID NO: 6);

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
  • Porcine circovivus loop-mediated isothermal amplification kit and application thereof
  • Porcine circovivus loop-mediated isothermal amplification kit and application thereof
  • Porcine circovivus loop-mediated isothermal amplification kit and application thereof

Examples

Experimental program
Comparison scheme
Effect test

Embodiment 1

[0078] The specificity result of embodiment 1 LAMP detection method

[0079] LAMP amplification was performed on 1 strain of porcine circovirus type 2, 7 strains of control virus and water control, and the results were as follows figure 1 As shown, the porcine circovirus type 2 reaction tube showed a rising curve of turbidity in about 20 minutes, which was a positive result, and the 7-strain control virus reaction tube and the water control reaction tube had no amplification, which was a negative result.

Embodiment 2

[0080] Example 2 Sensitivity results of LAMP detection method

[0081] The starting concentration of porcine circovirus type 2 genomic DNA was 1.27×10 2 ng / μL, after 10-fold serial dilution, carry out LAMP and common PCR amplification, the results are as follows figure 2 with image 3 As shown, the result shows that the detection limit of the LAMP method of the present invention is about 1.27 × 10 -3 ng / μL, while the detection limit of common PCR method is 1.27×10 -2 ng / μL.

Embodiment 3

[0082] Example 3 Fluorescence visualization detection results of LAMP detection method

[0083] According to the optimized conditions monitored by the turbidimeter, fluorescent dyes were added, reacted at 63°C for 60 minutes, and observed under ultraviolet light. Figure 4 To observe the results, the left tube is the reaction with porcine circovirus type 2 DNA as the template, which is a positive result, and the right tube is a negative control, which is a negative result. The test results show that the established LAMP method can be used conveniently at the grassroots level. You only need to use the kit with the LAMP primer designed by this method, add the sample, and use an inexpensive water bath to keep it at 63°C for 60 minutes to quickly observe the results without the need Open the lid to avoid contamination.

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 a porcine circovivus loop-mediated isothermal amplification (LAMP) kit and application thereof. The kit comprises LAMP primers, a 2* reaction buffer solution, a Bst DNA polymerase, a fluorescent visual detection reagent, ultrapure water and a porcine circovivus DNA template, wherein the LAMP primers comprises outer primers F3 and B3, inner primers FIP and BIP and loop primers LF and LB. Specificity detection and sensitivity detection prove that, the LAMP kit can be used for monitoring the reaction in real time, quantitatively detecting the copy number of porcine circovivuses and quickly and accurately obtaining the detection result, and brings convenience for simply, quickly and reliably detecting the porcine circovivuses.

Description

technical field [0001] The invention relates to the technical field of microbial detection, in particular to a fast, visualized and real-time quantitative detection of porcine circovirus ring-mediated isothermal amplification kit and its application. Background technique [0002] Circoviruses (PCVs) belong to the Circoviridae family and are the smallest known vertebrate viruses. Pigs are the main host of circovirus, and pigs of all ages have strong susceptibility to circovirus. Pigs infected during the embryonic period or early postnatal period often develop the disease after weaning, generally concentrated at the age of 5 to 18 weeks, especially at the age of 6 to 12 weeks. Pregnant sows infected with circovirus lead to reproductive impairment and can be vertically transmitted to piglets through the placenta. Infected pigs can excrete the virus from nasal fluid, feces and other excreta, and cause infection through the digestive tract and respiratory tract. [0003] A ne...

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
Patent Type & Authority Applications(China)
IPC IPC(8): C12Q1/70C12Q1/68C12R1/93
CPCC12Q1/701C12Q1/6844C12Q1/70C12Q2531/119
Inventor 彭昊李军冯世文潘艳陈泽祥胡帅杨威钟舒红谢宇舟禤雄标马春霞陶立柳锋谢永平许力干韦志锋秦若甫兰美益
Owner GUANGXI VETERINARY RES INST
Who we serve
  • R&D Engineer
  • R&D Manager
  • IP Professional
Why Eureka
  • Industry Leading Data Capabilities
  • Powerful AI technology
  • Patent DNA Extraction
Social media
Try Eureka
PatSnap group products