江苏农业学报
江囌農業學報
강소농업학보
JIANGSU JOURNAL OF AGRICULTURAL SCIENCES
2015年
3期
613-618
,共6页
胡成%朱善元%左伟勇%王安平%吴双%洪伟鸣%成大荣%马丽丽%王永娟
鬍成%硃善元%左偉勇%王安平%吳雙%洪偉鳴%成大榮%馬麗麗%王永娟
호성%주선원%좌위용%왕안평%오쌍%홍위명%성대영%마려려%왕영연
鸭I型肝炎病毒%环介导等温扩增%可视化检测
鴨I型肝炎病毒%環介導等溫擴增%可視化檢測
압I형간염병독%배개도등온확증%가시화검측
duck hepatitis virus type I%reverse transcription loop-mediated isothermal amplification
本研究根据GenBank中登录的鸭I型肝炎病毒( DHV I) VP1基因的高度保守序列,设计了特异性的引物,建立了一种灵敏、特异、高效的可视化体外环介导等温扩增方法( RT-LAMP)。结果表明,该方法的灵敏性可达到10 fg,是常规一步法RT-PCR方法的100倍以上。全部反应可以在1.0~1.5 h内完成,并可以直接通过肉眼观察颜色进行结果判定,该方法对其他鸭常见的病原体检测结果全部为阴性,该方法的建立为下一步研制鸭I型肝炎病毒的RT-LAMP快速检测试剂盒打下基础。
本研究根據GenBank中登錄的鴨I型肝炎病毒( DHV I) VP1基因的高度保守序列,設計瞭特異性的引物,建立瞭一種靈敏、特異、高效的可視化體外環介導等溫擴增方法( RT-LAMP)。結果錶明,該方法的靈敏性可達到10 fg,是常規一步法RT-PCR方法的100倍以上。全部反應可以在1.0~1.5 h內完成,併可以直接通過肉眼觀察顏色進行結果判定,該方法對其他鴨常見的病原體檢測結果全部為陰性,該方法的建立為下一步研製鴨I型肝炎病毒的RT-LAMP快速檢測試劑盒打下基礎。
본연구근거GenBank중등록적압I형간염병독( DHV I) VP1기인적고도보수서렬,설계료특이성적인물,건립료일충령민、특이、고효적가시화체외배개도등온확증방법( RT-LAMP)。결과표명,해방법적령민성가체도10 fg,시상규일보법RT-PCR방법적100배이상。전부반응가이재1.0~1.5 h내완성,병가이직접통과육안관찰안색진행결과판정,해방법대기타압상견적병원체검측결과전부위음성,해방법적건립위하일보연제압I형간염병독적RT-LAMP쾌속검측시제합타하기출。
Based on the sequences of duck hepatitis virus type I ( DHV I) VP1 gene available in GenBank, a series of specific primers were designed and reverse transcription loop-mediated isothermal amplification ( RT-LAMP) was devel-oped for visualized detection of DHV I . The results showed that the detection limit of the DHV I by RT-LAMP was 10 fg, 100 time higher than that by normal one-step RT-PCR. The amplification could be finished within 1. 0-1. 5 h e-valuated by naked eyes. The RT-LAMP developed in this study was specific and had no cross-reactivity with other pathogens of duck, laying a foundation for the development of RT-LAMP-based rapid detection kit.
