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《基于耐輻射球菌的生物傳感器的構(gòu)建以及耐輻射球菌drasnf2基因的研究》由會員上傳分享,免費在線閱讀,更多相關內(nèi)容在學術論文-天天文庫。
1、突變株,經(jīng)過DNA損傷實驗,發(fā)現(xiàn)draSNF2對耐輻射球菌的極端抗輻射特性并不是必需的。VllAbstractBasedonageneticallymodifiedradioresistantbacteriaDeinococcusradiodurans,weconstructedarealtimewholecellbiosensortomonitorradioactivityandgenotoxicityinhi曲Iyradioactiveenvironment。Theerahancedgreenfluorescenceprotein(eGFP)wasfusedtothepromoter
2、ofacrucialDNAdamage—induciblerecAgenefrom/9.radiodurans,andtheconsequentDNAfragment(PrecA-eg茲o)carriedbyplasmidwasintroducedintonradioduransR1straintoobtainthebiosensorstrainDRG300.ThisengineeredstrainCallexpresseGFPproteinandgeneratefluorescenceininductionoftherecAgenepromoter.Basedonthecorrelat
3、ionbetweenfluorescenceintensityandproteinexpressionlevelinliveD:radioduranscells,wediscoveredthatthefluorescenceinductionofstrainDRG300respondsinaremarkabledose-dependentmannerwhentreated誠m轉(zhuǎn)NAdamagesourcessuchasgammaradiationandmitomycinC。Itisencouragingtofindthewidelydetectiverangeandhi曲sensitiv
4、ityofthisreconstructedstraincomparing、蕊也otherwholecellbiosensorsinformerreports.TheseresultssuggestthestrainDRG300apotentialwholecellbiosensortoconstructadetectivesystemtomonitorthebiolo西calhazardsofradioactiveandtoxicpollutantsinenvironmentinrealtime.弧eBeta-Glo癬AssaySystemisahomogeneousmethodofq
5、uantitating13-galactosidaseexpressioninmammaliancells+強esystemprovidesabrightluminescentsignalthatisstableoverseveralhoursincommonlyusedcellculturemediumwithoutpriorsampleprocessing.Thehomogeneousassayprocedureinvolvestheadditionofasinglereagentdirectlytocellsculturedinserum-supplementedmedium。Th
6、roughputratesofseveralthousandsamplesperhourmaybeachieved謝也hi曲reproducibilityunderstandardlaboratoryconditions.WiththisIleWtechnology,weconstructedanewsystemforwholeeellbiosensor.HomologsofSnf2familymemberin疊radioduransR1,draSNF2WassequencingedtodefineasingleORFincludingtheon-linetagDRl258andDRl2
7、59。ThenewdefineddraSNF2genewithouttheinteinsequenceWasanalyzedwithSSO1653,andpredicteditstructureandbiologicalfunction.StudybytreatingDradioduransRlstrainsanddraSNF2function—deficientmutantwithionizingradiationandMMCwe