Development of a Loop-Mediated Isothermal Amplification Assay for Porcine Circovirus Type 2

• Published in: Virologica Sinica Vol. 26; no. 3; pp. 214 - 220
• Main Authors: Liu, Ye-bing, Zhang, Lei, Xue, Qin-hong, Ning, Yi-bao, Zhang, Zhi-gang
• Format: Journal Article
• Language: English
• Published: Berlin/Heidelberg Springer-Verlag 01.06.2011; China Institute of Veterinary Drug Control,Being 100081,China%China Institute of Veterinary Drug Control,Being 100081,China; College of Veterinary Medicine,Northwest Agriculture & Forestry University,Yangling 712100,China
• Subjects: Animals; Biochemistry; Biomedical and Life Sciences; Biomedicine; Circoviridae Infections; Circoviridae Infections - diagnosis; Circoviridae Infections - veterinary; Circoviridae Infections - virology; Circovirus; Circovirus - genetics; Circovirus - isolation & purification; conserved sequences; detection limit; diagnosis; DNA; DNA Primers; DNA Primers - genetics; DNA模板; GenBank; genetics; isolation & purification; Medical Microbiology; methods; Microbial Genetics and Genomics; Microbiology; Nucleic Acid Amplification Techniques; Nucleic Acid Amplification Techniques - methods; Oncology; PCV2; polymerase chain reaction; Porcine circovirus; Porcine circovirus-2; Swine; Swine Diseases; Swine Diseases - diagnosis; Swine Diseases - virology; veterinary; Virology; viruses; 介导; 实验; 检测系统; 猪圆环病毒2型; 等温; Loop-mediated isothermal amplification (LAMP); Porcine circovirus type 2 (PCV2); Virus detection; Porcine circovirus type 2(PCV2); Loop-mediated isothermal amplification(LAMP)
• ISSN: 1674-0769; 1995-820X; 1995-820X
• DOI: 10.1007/s12250-011-3169-x

In this study, the loop-mediated isothermal amplification （LAMP） method was used to develop a rapid and simple detection system for porcine circovirus type 2 （PCV2）. According to the PCV2 sequences published in GenBank, multiple LAMP primers were designed targeting conserved sequences of PCV2. Using the DNA extracted from PCV2 isolates HUN-09 and SD-09 as the template, LAMP reactions in a PCV2 LAMP system was performed, the amplification products were detected by adding SYBR Green I and could be observed directly by the naked eye. The results showed highly-efficient and specific amplification in 30 min at 63°C with a LAMP real-time turbidimeter. Furthermore, PCV2 DNA templates, with a detection limit of 5.5×10–5 ng of nucleic acid, indicated that this assay was highly sensitive. The results obtained with the naked eye after SYBR Green I staining were consistent with those detected by the real-time turbidimeter, showing the potential simplicity of interpretation of the assay results. The LAMP assay appeared to have greater accuracy than PCR and virus isolation for the analysis of 18 clinical samples. In addition it offers higher specificity and sensitivity, shorter reaction times and simpler procedures than the currently available methods of PCV2 detection. It is therefore a promising tool for the effective and efficient detection of PCV2.