Description:
STAT3ResponsiveLuciferaseReporterHelaStableCellLineisderivedfromhumancervicalcancer,andstablyexpressfireflyluciferasereportergeneunderthecontroloftheSTAT3responseelement. ThiscelllineisanidealcellularmodelformonitoringtheactivationofJAK-STATReceptorSignalingPathwaytriggeredbystimulitreatment,enforcedgeneexpressionandgeneknockdown.Principle:
STAT3isactivatedinresponsetovariouscytokinesandgrowthfactorsincludingIFNγ,IL-6andOSM.TheactivationofSTAT3resultsinformationofSTAT3homodynesandSTAT3/STAT1heterodimersthattranslocatetothecellnucleusandinducetranscriptionofgenesbybindingtotheconsensuselementonthepromoterregionoftargetgenesassociatedwithcellgrowthandapoptosis.SignosishasestablishedStat3luciferasereporterHeLastablecellline,whichcanbeused asareportersystemformonitoringtheactivationofStat3triggeredbystimulitreatment,enforcedgeneexpressionandgeneknockdown.
Thecelllinewasestablishedbytransfection usinga pTA-Stat3-luciferasereportervector,whichcontains4repeatsofStat3bindingsites,aminimalpromoterupstreamofthefireflyluciferasecodingregion, alongwithhygromycinexpressionvectorfollowedbyhygromycinselection. Thehygromycinresistantclonesweresubsequentlyscreenedforoncostatininducedluciferaseactivity.Theclonewiththehighestfoldinduction(10fold)wasselectedandexpandedtoproducethisstablecellline.
PrinciplebehindTFluciferasereporter. TFluciferasereporterstablecelllineutilizesartificialpromoterconstructstodriveluciferaseexpression. Thepromoterregioncanconsistsofmultiplerepeatsofacis-elementTFbindingsite,aDNAfragmentfromthepromoterregionofaknownTFdownstreamgene,oraDNAfragmentcontainingputative/knownTFbindingsites. ThereareseveralwaysthataTFcanbeactivated,suchasthroughextracellularstimuliorthroughintracellularsignalingpathways. Onceactivated,theTFtranslocatestothenucleusandofteninteractswithrelevantco-factorstodrivegeneexpression. Onceluciferaseisexpressed,itcangeneratelightinanenzymaticassayandtheamountoflightmeasuredispositivelycorrelatedwiththelevelofTFactivation. |
Data:
Figure1:AnalysisofHeLa/STAT3-luciferaseactivityofcontrolandtreatmentwithOncostatin. Thecellswereseededona96-wellplateforovernightwithDMEMincluding10%FBS.Thecellsthenweretreatedwithorwithout10ng/mlOncostatinrespectivelyinDMEMand0.1%FBSfor8hours. Stat3luciferasereportercelllineshowedmorethan25-foldincreaseinluciferaseactivityinresponsetoOncostatinMtreatment.
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至于到底好不好转就跟你的细胞有关了，和细胞是否是稳转株关系不大。
推荐了解“主细胞库”“工作细胞库”这两个名词。
推荐了解“主细胞库”“工作细胞库”这两个名词。
如果经费充足的话可以找公司包装病毒，如武汉的普健可以提供各种载体的构建及细胞株构建的技术服务。
然后你得确定，这个细胞株对于你得研究来说不可或缺，具有明确的代表性。
上面两个都没有问题的话，祝你顺利。
