TY - JOUR
T1 - Development of one-step reverse transcription loop-mediated isothermal amplification (LAMP) as a screening tool for influenza A viruses
AU - Tun, Hein Min
AU - Wisedchanwet, Trong
AU - Wongphatcharachai, Manoosak
AU - Nonthabenjawan, Nutthawan
AU - Chaiyawong, Supasama
AU - Theamboonlers, Apiradee
AU - Poovorawan, Yong
AU - Amonsin, Alongkorn
PY - 2014
Y1 - 2014
N2 - 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 63oC for 60 min 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.
AB - 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 63oC for 60 min 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.
KW - Influenza A
KW - Reverse transcription loop-mediated isothermal amplification
UR - http://www.scopus.com/inward/record.url?scp=84929300469&partnerID=8YFLogxK
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M3 - Article
AN - SCOPUS:84929300469
SN - 0125-6491
VL - 44
SP - 427
EP - 434
JO - Thai Journal of Veterinary Medicine
JF - Thai Journal of Veterinary Medicine
IS - 4
ER -