Development of One-Step Reverse Transcription Loop-Mediated Isothermal Amplification (LAMP) as a Screening tool for Influenza A Viruses

作者：Tun Hein Min; Wisedchanwet Trong; Wongphatcharachai Manoosak; Nonthabenjawan Nutthawan; Chaiyawong Supasama; Theamboonlers Apiradee; Poovorawan Yong; Amonsin Alongkorn^*

来源：Thai Journal of Veterinary Medicine, 2014, 44(4): 427-434.

• Influenza A
• reverse transcription loop-mediated isothermal amplification

Influenza A virus is a major cause of influenza pandemics and can infect several host species including humans and animals. The objective of this study was to develop a one-step reverse transcription loop-mediated isothermal amplification assay (LAMP) for the detection of genetically diverse influenza A viruses from both human and animal hosts. First, a set of two inner and two outer primers were designed based on the conserved region of the matrix (M) gene of influenza A viruses. The amplification reaction was optimized at 63 degrees C for 60 mm and performed in a simple heat block. The amplicons could be visualized either by gel electrophoresis or by visual analysis upon addition of SybrGreen. The developed LAMP assay was tested with 50 influenza A isolates including H1N1, H1N2, H3N2, H5N1 and H7N4 from swine, avian and human hosts. In sensitivity test, the assay detection capability was ten times more sensitive than conventional RT-PCR and comparable to real time RT-PCR. In summary, this assay is a rapid, simple and sensitive assay suitable for less-equipped laboratories and thus can be utilized in the field as a screening test.

• 出版日期2014-12