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Development of a direct duplex real-time PCR assay for rapid testing of domestic cat hepadnavirus

##article.authors##

  • Maya Shofa Department of Veterinary Science, Faculty of Agriculture, University of Miyazaki
  • Akiho Ohkawa Department of Veterinary Science, Faculty of Agriculture, University of Miyazaki
  • Tamaki Okabayashi Department of Veterinary Science, Faculty of Agriculture, University of Miyazaki
  • Yasuyuki Kaneko Veterinary Teaching Hospital, Faculty of Agriculture, University of Miyazaki
  • Saito, Akatsuki Department of Veterinary Science, Faculty of Agriculture, University of Miyazaki

DOI:

https://doi.org/10.51094/jxiv.112

キーワード:

domestic cat hepadnavirus、 direct duplex real-time PCR assay、 rapid、 sensitive

抄録

Domestic cat hepadnavirus (DCH) is a novel hepadnavirus, first identified in 2018. The DCH is generally diagnosed using conventional polymerase chain reaction (PCR) assays, which include a time-consuming agarose gel electrophoresis. In this study, we developed a rapid, sensitive, and specific real-time PCR assay for the detection of the DCH genome. To streamline the procedure, our real-time PCR assay was carried out using blood samples, without deoxyribonucleic acid (DNA) extraction. A primers/probe set was designed based on the nucleotide sequences of the surface gene of the DCH strain Japan/KT116/2021 (Accession# LC668427), which we recently identified from a feline blood sample in Japan. To exclude the possibility that the PCR reaction was blocked by anticoagulants, we also used a primers/probe set for amplifying the housekeeping beta-actin gene. Direct duplex real-time PCR assay had a high sensitivity, with a limit of detection of 10 copies/μL for DCH. We successfully established a rapid and highly sensitive duplex real-time PCR assay for the detection and quantification of DCH. This direct duplex real-time PCR assay is a useful tool for DCH diagnosis and surveillance.

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引用文献

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投稿日時: 2022-07-07 03:14:52 UTC

公開日時: 2022-07-07 08:55:49 UTC
研究分野
生物学・生命科学・基礎医学