Ou Shan-Chia, Giambrone Joseph J, Macklin Kenneth S
Poultry Science Department, Auburn University, Auburn, AL 36849-5416, USA.
J Vet Diagn Invest. 2012 Jan;24(1):138-41. doi: 10.1177/1040638711427578. Epub 2011 Dec 6.
A TaqMan real-time polymerase chain reaction (PCR) and loop-mediated isothermal amplification (LAMP) assay were developed to detect Gallid herpesvirus 1 (GaHV-1, formerly Infectious laryngotracheitis virus). The standard curve of real-time PCR was established, and the sensitivity reached 10 copies/μl. In the current study, the conversion between viral titer and GaHV-1 genomic copy number was constructed. Six primers for LAMP assay amplified target gene at 65°C within 45 min, and the detection limit was 60 copies/μl. The 6 primers were highly specific, sensitive, and reproducible for detection of GaHV-1. Although the sensitivity of LAMP was lower than that of real-time PCR, LAMP was faster, less expensive, and did not require a thermocycler. The LAMP assay would be a viable alternative assay in diagnostic laboratories that do not employ real-time PCR technology.
开发了一种TaqMan实时聚合酶链反应（PCR）和环介导等温扩增（LAMP）检测方法，用于检测鸡疱疹病毒1型（GaHV-1，以前称为传染性喉气管炎病毒）。建立了实时PCR的标准曲线，灵敏度达到10拷贝/μl。在本研究中，构建了病毒滴度与GaHV-1基因组拷贝数之间的转换关系。用于LAMP检测的6条引物在65°C下45分钟内扩增靶基因，检测限为60拷贝/μl。这6条引物对GaHV-1的检测具有高度特异性、敏感性和可重复性。虽然LAMP的灵敏度低于实时PCR，但LAMP速度更快、成本更低，且不需要热循环仪。LAMP检测方法对于不采用实时PCR技术的诊断实验室将是一种可行的替代检测方法。
