简介:目的:构建40S核糖体蛋白S6的原核表达载体,表达并纯化S6蛋白,将其作为底物用于S6激酶(S6K)的体外活性测定。方法:采用RT-PCR方法从人胚肾细胞HEK293中获取S6cDNA,将扩增产物克隆至大肠杆菌表达载体中,进行酶切及测序鉴定;IPTG诱导GST-S6融合蛋白在大肠杆菌中表达,用谷胱甘肽亲和层析纯化GST-S6,免疫沉淀法检测该蛋白是否可作为底物用于S6K的体外激酶活性测定。结果:酶切及测序鉴定表明构建了S6原核表达载体,并表达及纯化出GST-S6融合蛋白,相对分子质量为55×103。该蛋白可用于S6K的体外激酶活性测定,特异性强。结论:S6蛋白的克隆、表达与纯化成功,可用于S6K的体外激酶活性测定,为研究S6K的功能奠定了基础。
简介:预防与控制乙型肝炎发病的乙型肝炎病毒(HBV)疫苗,是有重大的社会和经济意义。HBV的持续感染可引起慢性肝脏疾患,并逐步发展为肝硬化和肝细胞癌(HCC)。目前的乙肝重组亚单位疫苗可以使90%的接种者产生保护性抗体;但是对慢性HBV携带者,由于其机体对HBsAg蛋白产生耐受,不能产生体液和细胞免疫,因此它只能作为一种预防性的疫苗。DNA疫苗(基因疫苗)是一种新的疫苗技术,通过向体内递送编码抗原的细菌质粒,刺激产生特异的体液和细胞免疫反应。在小鼠和其他的肝炎病毒感染动物模型中,HBVDNA疫苗可以特异性地引起体液和细胞免疫,清除HBV转基因动物血循环中的HBsAg颗粒和HBVDNA。如果加入各种免疫调节细胞因子的基因,可以进一步提高HBVDNA疫苗的免疫效果,因此它不仅可作为预防性疫苗,也可作为治疗型疫苗。
简介:Themodulationandcontrolofgecko'sfootmovementswerestudiedelectrophysiologicallyinordertodesignthemotorcontrolsystemofagecko-mimicrobot.Inthisstudy(1)theanatomyoftheperipheralnervescontrollingthegecko'sfootmovementswasdetermined;(2)therelationshipbetweenthelimbnervesofthegeckoanditsfootmotorpatternswasstudied;(3)theafferentimpulsesofthenervesevokedbyrubbingthegecko'stoesandpalmwererecorded;(4)copyingthenaturalpatternsofmovementofthegecko'sfoot(abduction,adduction,flexion,andrevolution)anditslimbnervemodulationandcontrolmechanism,thenerveswerestimulatedundercomputercontrol,andtheresultsrecordedbyCCD.Resultssuggestthatgecko'sfootmovementscanbesuccessfullycontrolledbyartificialelectricalsignals.
简介:Thesandfishisalizardhavingtheremarkableabilitytomoveindesertsandinaswimming-likefashion.Themostout-standingadaptationstothismodeoflifearethelowfrictionbehaviourandtheextensiveabrasionresistanceofthesandfishskinagainstsand,outperformingevensteel.Weinvestigatedthetopography,thecompositionandthemechanicalpropertiesofsandfishscales.Theseconsistofglycosylatedkeratinswithhighamountofsulphurbutnohardinorganicmaterial,suchassilicatesorlime.Remarkably,atomicforcemicroscopyshowsanalmostcompleteabsenceofattractiveforcesbetweenthescalesurfaceandasilicontip,suggestingthatthisisresponsiblefortheunusualtribologicalproperties.Theunusualglycosylationofthekeratinswasfoundtobeabsolutelynecessaryforthedescribedphenomenon.Thescalesweredissolvedandreconstitutedonapolymersurfaceresultinginpropertiessimilartotheoriginalscale.Thus,weprovideapathwaytowardsexploitationofthereconstitutedscalematerialforfutureengineeringapplications.
简介:Naturalcomposites,formedthroughbiomineralization,havehighlyorderedstructureswhichhavebeenaptlyexploredforfunctionalapplications.Thoughtheroleoforganicphaseshasbeenwellunderstoodinbiomineralization,notenoughattentionhasbeenpaidtotheroleofbio-membraneswhichareoftenfoundencapsulatingthechamberinwhichmineralizationoccurs.Wehaveusedthenaturalproteinandsemi-permeablemembraneofchickeneggstogrowdifferentmaterialssuchasceramics,semi-metalsandmetalstounderstandtheroleofbio-membranesinbiomineralization.Weherereportthesuccessfulbiomimeticsynthesisofcalcite,cadmiumsulphide,andsilverhavinghomogeneousmorphologies.Wehavefoundthatthemembraneoperateslikeatunedgateway,playingasignificantroleincontrollingthemorphologyoftheinorganiccrystalsformedduringbiomineralization.
简介:目的:建立无线粒体DNA(mtDNA)的人肺腺癌ρ^0A549细胞系。方法:在含50ng/mL溴化乙锭(EB)、100μg/mL丙酮酸钠和50μg/mL尿嘧啶核苷的RPMI1640细胞培养基中传代培养A549细胞;用低剂量EB连续诱导培养35d后,采用光镜观察、TaqMan探针法实时荧光定量PCR(qPCR)和Western印迹鉴定无mtDNA的ρ^0A549细胞系;采用MTT法测定ρ^0A549细胞增殖曲线。结果:倒置显微镜下野生型ρ^+A549细胞为多角形,ρ^0A549细胞形态呈拉长枝状;qPCR结果显示,低剂量EB诱导35d的ρ^0A549细胞中无mtDNA的存在。Western印迹结果显示,ρ^+A549细胞中能表达核基因编码的线粒体蛋白SDHA和ATP5A,也能表达线粒体基因组编码的蛋白MT-COXI和MT-ATP6;ρ^0A549细胞中无MT-COXI和MT-ATP6蛋白表达,但核基因编码的SDHA和ATP5A蛋白能够正常表达。MTT结果显示,与ρ^+A549细胞相比,ρ^0A549细胞生长速度明显减慢,差异有统计学意义(P<0.05)。结论:构建和鉴定了无mtDNA的人肺腺癌ρ^0A549细胞系,为后续探讨mtDNA缺失或突变与人肺腺癌发生之间的关系奠定了实验基础。