多重分子信标探针芯片构建与结核杆菌耐药基因多重突变位点检测研究

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1、第三军医大学博士学位论文DesignfortheChipLabelledwithMulti·molecularBeaconProbeandtheExplorationonMulti-sitesDetectionofBacillusTuberculosisAbstractobjective：ThecurrentdetectingmethodsforBacillusTuberculosis(TB)includemanytechniquessuchasX—ray,sputumsmear，cellcul

2、ture，medicinesensitivity，PCR，andSOon．Theyaretediousortime—consumingmethodswithlowsensitivityandspecificity,whichmayeasyleadtomisdiagnosisandeconomicoverburdenofpatients．AgreatvarietyofdrugsprescribedtopatientswithTuberculosisandtheiroveruseleadtothei

3、ncreasinglycomplicatedmutationofdrug—resistantgeneswithwidecomplexrangeofmutationsitesfromthesingle—basemutation，deletion，insertion，malpositiontothatwithmultiple—sitesandlargefragmentwhichpresentabarriertodetectingthedrugresistantmutationgenesaccurat

4、elyandcomprehensively．Especiallyforthetestforwide—existingsingle—basemutationsites，thetraditionaltechniquesincludingthetraditionalgenechipcanhardlyaddresstheproblems．Themulti—drug—resistanceisthemainfactoraffectingitstreatmentandpopularityforitssimpl

5、eindicatorandscarceinformationondrug—resistance．ThustoconstructasystemforidentifyingtheTBandtestingitdrugresistanceinafast，sensitive，comprehensiveandsimplewayisflissueurgenttoaddressedinthefieldofLaboratoryDiagnosticsbothathomeandabroad．Themolecularb

6、eaconprobeisanucleicacidprobewithgreatcapacityoftestingsingle—basemutationsites，however，duetoitsmolecularlimitation，therestillisblankinitspracticeapplicationinthefluorescencechips．Inthisstudy，weattempttoconstructachiptechniquelabelledbyMBofTBmultiple

7、resistancegenesmutationbymakingadvantageofthestrongerspecificitytodetectionofthesingle—basemutationsitesoftargetmolecularandthebeaconprobe，combiningwiththehighthroughputoffluorescence，whichiscapableofdirectlydetectingthespecificityofmultipletargetsit

8、esofdrugresistantgenesandthenucleicacidofTBandprovidesanewmethodtoidentifythedrugresistantgenesofTBandtomaketheepidemicinvestigation．MethodsandMaterials：1．BasedontheIS986genesequenceofTBfoundbyGenBank，wedesignedthe第三军医大学博士学位论文primersonthesoftwarePrim