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《水貂阿留申病毒、腸炎病毒與犬瘟熱病毒多重pcr檢測方法的建立與應(yīng)用》由會員上傳分享,免費(fèi)在線閱讀,更多相關(guān)內(nèi)容在應(yīng)用文檔-天天文庫。
1、水貂阿留申病毒、腸炎病毒與犬瘟熱病毒多重PCR檢測方法的建立與應(yīng)用馬芹王元智閏文卓趙麗麗陳洪巖陸濤峰物與比較醫(yī)學(xué)重點(diǎn)實(shí)驗(yàn)室目的水貂阿留中病、病毒性腸炎與犬瘟熱并稱影響水貂健康的三大疫病,擬建立一種可同時檢測三種病毒的多重PCR檢測方法。方法針對三種病毒基因保守區(qū)分別設(shè)計了3對特異性引物,對貂阿留申病毒(ADV)和水貂腸炎病毒(MEV)的DNA模板和犬瘟熱病毒(CDV)的RNA模板進(jìn)行丫多重PCR擴(kuò)增和擴(kuò)增條件優(yōu)化。結(jié)果PCR可同時擴(kuò)增出601bp(ADV)、205bp(MEV)和451bp(CDV)的特異性目的條帶,敏感性試驗(yàn)表明最低核酸檢出量為ADV每微升2.67X10
2、4拷貝、1EV每微升3.02X101拷貝和CDV每微升1.72X105拷貝。臨床樣品檢測結(jié)果表明多重PCR和單一PCR檢測結(jié)果-致。結(jié)論建立的多重PCR檢測方法可快速地檢測ADV、MEV和CDV單一或混合感染的臨床樣品。關(guān)鍵詞:多重PCR;水貂阿留中病毒;水貂腸炎病毐;犬瘟熱病毒;基金:國家支撐計劃項(xiàng)目(2015BA107B02)EstablishmentandapplicationofamultiplexPCRassayfordetectionofAleutiandiseaseparvovirus,enteritisparvovirusandcaninedistemp
3、ervirusintheminkMAQinWANGYuan-zhiYANWen-zhuoZHAOLi-liCHENHong-yanLUTao-fengHarbinVeterinaryResearchInstitute,ChineseAcademyofAgriculturalSciences,HeilongjiangProvincialKeyLaboratoryofLaboratoryAnimalandComprativeMedicine;Abstract:ObjectiveAleutiandisease,minkenteritisandcaninedistemperare
4、thethreemajordiseasesaffectinghealthofmink.ThisstudyintendstoestablishamultiplexPCRassayforsimultaneouslydetectingofthesethreeviruses.MethodsAccordingtotheconservativesequencesreportedinGenBank,threepairsofspecificprimersweredesignedtoamplifytheDNAtemplatesofAleutianminkdiseaseparvovirus(
5、ADV),minkenteritisparvovirus(MEV),andRNAtemplatesofcaninedistempervirus(CDV),andoptimizedtheamplifyingconditions.ResultsThespecificobjectivestripsof601bp(ADV),205bp(MEV)and451bp(CDV)wereamplifiedsimultaneously.Thesensitivitytestshowedthatthelowestnucleicaciddetectionlimitswere2.67X101copi
6、esperpLforADV,3.02X101copiesperPLforMEV,and1.72X10)copiesperPLforCDV.TheresultsoftestoftheclinicalsamplesshowedthatthemultiplePCRandsinglePCRassaywereconsistent.ConclusionsTheestablishedmultiplexPCRassayinthisstudycanbeusedtorapidlydetecttheclinicalsamplesofADV,MEVandCDVsingleormixedinfec
7、tions.Keyword:MultiplexPCR;Aleutianminkdiseaseparvovirus;Minkenteritisparvovirus;Caninedistempervirus;貂作為重要的經(jīng)濟(jì)動物,不僅僅具有經(jīng)濟(jì)價值U1,同時貂還是流感病毒、SARS、新城疫病毒、貂圓環(huán)病毒、阿留申病毒以及腸炎病毒的宿主,其中雪貂對流感病毒十分易感,感染后表現(xiàn)出與人相似的癥狀,還能產(chǎn)生免疫反應(yīng),因此雪貂可作為研究流感病毒最佳的實(shí)驗(yàn)動物模型。貂的實(shí)驗(yàn)動物化養(yǎng)殖必須保證背景淸晰,并對其攜帶的病原體進(jìn)行控制,所以需要建立一