• Title/Summary/Keyword: Ultra-rapid nested PCR

Search Result 2, Processing Time 0.016 seconds

Development of Ultra-rapid Nested PCR Method for Detection of Specific Gene of Tracheal Mite (Acarapis woodi) (기문응애(Acarapis woodi) 특이 유전자 검출을 위한 초고속 nested PCR법 개발)

  • Kim, MoonJung;Kim, Byoung-Hee;Kim, SoMin;Truong, A Tai;Kim, Jung-Min;Kim, Seonmi;Yoon, Byoung-Su
    • Journal of Apiculture
    • /
    • v.34 no.1
    • /
    • pp.15-26
    • /
    • 2019
  • Tracheal mite (Acarapis woodi) is an internal parasite that is parasitic on the bronchus of adult bees and sucks fluid from the trachea. Since its first report by Rennie, it has been spread throughout Europe and in some Asian regions, with adjacent Japan and China reported in 2011 and 2012, respectively. Korea detected specific genes of A. woodi in 2015, but only one of 99 samples has been identified and the being of A. woodi has not been confirmed. In this study, we established a specific nested PCR method to confirm for detecting low-copy number of A. woodi-specific gene in bee samples. As a result, A. woodi-specific COI gene was amplified in 15 of 23 samples, and they were judged positive by melting point analysis and sequencing analysis. Although we could not observe the existence of the mites in bees, our results suggest that tracheal mit might exist in nature.

Quantitative Detection of Tropilaelaps in Hive by Specific Gene Detection from Hive Debris (봉변에서 특이 유전자 검출법에 의한 봉군 내 꿀벌가시응애류 (Tropilaelaps)의 정량적 검출)

  • Kim, Byounghee;Kim, Somin;Kim, Moonjung;Kim, Jungmin;Truong, A Tai;Kim, Seonmi;Yoon, Byoungsu
    • Journal of Apiculture
    • /
    • v.34 no.1
    • /
    • pp.27-37
    • /
    • 2019
  • Rapid detection of Tropilaelaps, an external parasite of honeybees that lead to malformation of honeybee or colony collapse disorder, is becoming important. But it is very difficult to find with the naked eye of Tropilaelaps. In this study, we have developed a method to detect the specific gene of Tropilaelaps from the hive debris and to know the number of Tropilaelaps in the hive through Tropilaelaps-specific quantitative detection. Tropilaelaps-specific gene amplified in DNA extracted from hive debris by consecutive PCR (1st detection, 2nd nested PCR). It could detect 101 molecules level of Tropilaelaps-specific gene and confirm the amplification of the Tropilaelaps-specific gene. It was possible to accurately quantify the number of Tropilaelaps from the hive debris sample, which is difficult to discriminate the presence of Tropilaelaps visually, through Tropilaelaps-specific detection. Under the microscope, Tropilaelaps was collected and quantitative detection of Tropilaelaps-specific genes was performed. It was possible to quantify the number of Tropilaelaps present in the hive through the molecules of the quantified Tropilaelaps-specific genes. We suggest that hive debris can represent as a micro-environment to hive and show that it can be a simpler and more accurate sample than using a parasitic host honeybee. We expect that hive debris should facilitate the monitoring of Tropilaelaps in hive.