Cherry leaf roll virus의 신속 및 특이적 진단을 위한 등온증폭법 개발
Development of Diagnostic System for Rapid and Specific Detection of Cherry leaf roll virus
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초록
Cherry leaf roll virus(CLRV)는 group IV positive sense ssRNA viruses, Nepovirus로 분류되는식물병원성 바이러스이다. CLRV는 체리 등 목본 및 완두 등 콩과 작물을 자연 기주로 하며, 실험적으로약 36개 과 이상의 넓은 기주 범위를 가지고 있어 국가적, 경제적 및 농가 개인적 피해를 야기 할 가능성이 보고되고 있다. 현재 CLRV를 검출하는 방법으로 역전사(reverse transcription; RT)-nesdted 중합효소연쇄반응(polymerase chain reaction; PCR) 이 활용되고 있으며, 다양한 기주로부터 CLRV를검출하기 위해서는 검출 감도, 특이성, 반응 시간, 단순성 등이 중요 요소였다. 그러나 RT-nestedPCR은 두 단계로 구성되어 있어 단순하지 않고, CLRV를 검출하는데 약 10시간 이상의 반응 시간이 소요되었다. 이번 연구에서는 등온증폭법을 이용하여 단순하고 신속하게 CLRV를 검출하는 방법을 개발하였다. 등온증폭 반응은 RT-nested PCR과 동등한 검출 감도로 CLRV를 검출 하였다. 그러나 반응 시간을 약 2시간 수준으로 단축하였으며, 6개 영역을 사용하는 등온증폭 프라이머의 사용으로 더욱 특이적으로 증폭 할 수 있었다
Abstract
Cherry leaf roll virus(CLRV) is a plant virus of belonging to Nepovirus, group IV positivesense ssRNA viruses. CLRV has a wide host range, and it has been found in up to 36families, including cherry, woody plant and pea families. When related crops were infected byCLRV, it cause to significant national, economic and farm damage. Reverse transcription(RT)-nested polymerase chain reaction(PCR) is used as the major technique for the detectingCLRV from various hosts being that important of sensitivity, specificity, reaction time andsimple. However, two-steps nested PCR was not simple and need more ten hours reaction timefor detecting CLRV. For simple and rapid detection of CLRV, using the loop-mediatedisothermal amplification(LAMP) assay was developed, this study. In the results, LAMP reactionshowed similar sensitivity compare with RT-nested PCR. However, running time was reduced toapproximately two hours, and can more specific detection of CLRV using six regions LAMPprimers.
참고문헌 (18)
- 안영창, "등온 증폭법을 이용한 결핵균의 빠른 검출 시스템 개발", 대한화학회지, 52(3), pp.273~280 (2008), 대한화학회
- Buchhop J, "Differentiation of Cherry leaf roll virus isolates from various host plants by immunocapture-reverse transcription-polymerase chain reaction-restriction fragment length polymorphism according to phylogenetic relations", J. Virol. Methods, 157, pp.147~154 (2009)
- Caruso P, "A new and sensitive co-operational polymerase chain reaction for rapid detection of Ralstonia solanacearum in water", J. Microbiol. Meth, 55, pp.257~272 (2003)
- 조민호, "칩 기반 등온증폭법을 이용한 약제 내성 포도상구균의 검출", 대한화학회지, 57(1), pp.81~87 (2013), 대한화학회[DOI]
- Eastwell KC, "Complete nucleotide sequences and genome organization of a cherry isolate of Cherry leaf roll virus", Arch. Virol, 157, pp.761~764 (2012)
- Gallas-Lindemann C, "Giardia and Cryptosporidium spp. dissemination during wastewater treatment and comparative detection via immunofluorescence assay(IFA), nested polymerase chain reaction(nested PCR) and loop mediated isothermal amplification(LAMP)", Acta Trop, 6, pp.43~51 (2016)
- Cherry leaf roll virus, "Jones AT.", Association of Applied Biologists, ~, pp.AAB Descriptions of plant viruses No. 306 (1985)
- Kim JH, "Development of simple and rapid diagnostic method for Strawberry latent ring-spot virus in plants using loop-mediated isothermal amplification assay", J. Plant Pathol. Microbiol, 7, pp.377~ (2016)
- 이시원, "Loop-mediated Isothermal Amplification Assay to Rapidly Detect Wheat Streak Mosaic Virus in Quarantined Plants", The Plant Pathology Journal, 31(4), pp.438~440 (2015), 한국식물병리학회[DOI]
- 이시원, "종자전염검역바이러스 3종(CRLV, SpLV 및 WClMV)을 검정하기 위한 RT-PCR 및 Nested PCR 프라이머 개발", 농업생명과학연구, 48(3), pp.75~83 (2014), 농업생명과학연구원[DOI]
- 이시원, "RT-PCR과 nested PCR을 이용한 Nepovirus속 식물검역 바이러스 4종의 정밀진단", 식물병 연구, 19(3), pp.220~225 (2013), 한국식물병리학회
- Notomi T, "Loopmediated isothermal amplification of DNA", Nucleic Acids Res, 28, pp.E63~ (2000)
- Parida M, "Loop mediated isothermal amplification(LAMP): a new generation of innovative gene amplification technique; perspectives in clinical diagnosis of infectious diseases", Rev. Med. Virol, 18, pp.407~421 (2008)
- Priou S, "An improved enrichment broth for the sensitive detection of Ralstonia solanacearum(biovars 1 and 2A) in soil using DAS-ELISA", Plant Pathology J, 55, pp.36~45 (2006)
- Sanfaçon H, "Secoviridae: a proposed family of plant viruses within the order picornavirales that combines the families sequiviridae and comoviridae, the unassigned genera cheravirus and sadwavirus, and the proposed genus torradovirus", Arch. Virol, 154, pp.899~907 (2009)
- Sidoti F, "Alternative molecular tests for virological diagnosis", Mol. Biotechnol, 53, pp.352~362 (2013)
- National diagnostic protocol for Cherry leaf roll virus, CLRV(cherry and walnut strains), "SPHDS.", Australian Government Department of Agriculture, ~ (2016)
- Suwancharoen D, "A nationwide survey of pathogenic leptospires in urine of cattle and buffaloes by loop-mediated isothermal amplification (LAMP) method in Thailand, 2011-2013", J. Vet. Med. Sci, 78, pp.1495~1500 (2013)