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: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:研究肿瘤坏死因子受体相关因子6(TRAF6)对NOD样受体蛋白3(NLRP3)炎症小体信号通路调控作用的机制。方法:利用免疫共沉淀和免疫印迹在HEK-293T细胞中研究TRAF6与NLRP3的相互作用;通过检测乳酸脱氢酶(LDH),在THP-1细胞中研究TRAF6对NLRP3炎症小体信号通路活性的影响。结果:TRAF6通过与NLRP3的相互作用增加NLRP3的稳定性,进而促进NLRP3炎症小体信号通路介导的LDH的释放。结论:TRAF6通过增加NLRP3的稳定性正调控NLRP3炎症小体信号通路。
简介:目的:分离羽衣甘蓝S13-b位点受体激酶(SRK13-b)基因并进行序列及结构域分析,构建SRK13-b结构域的原核表达载体并进行重组蛋白质的原核表达和纯化。方法:提取羽衣甘蓝S13-bS13-b自交不亲和系花期柱头的RNA,用RT-PCR法分离SRK13-b基因;将编码SRK13-b激酶结构域的序列插入大肠杆菌表达载体pET-14b中,构建原核表达质粒pET-SRK13-bCT,转化大肠杆菌BL21(DE3)pLysS菌株,经0.1mmol/LIPTG诱导,用Ni—NTA亲和层析柱对SRK13-b激酶结构域蛋白进行纯化。结果:分离获得羽衣甘蓝SRK13-b基因的长度为2571bp,编码856个氨基酸,GenBank收录号为EU180597;对SRK13-b激酶结构域蛋白进行诱导表达及纯化,SDS—PAGE显示相对分子质量约43×10^3的蛋白质特异表达,对表达产物进行分离纯化,获得了SRK13-b激酶结构域的融合蛋白。结论:羽衣甘蓝SRK13-b基因的克隆及激酶结构域的原核表达,为研究SRK的功能及自交不亲和性奠定了基础。
简介:目的:构建一个牛dSl酪蛋白调控序列指导人溶菌酶(hLYZ)基因组序列的杂合基因座。方法:采用本实验室发明的连续3步缺口修复技术。将6个无痕连接的同源臂插入以pBR322为载体的骨架中,构成能进行3次连续基因抓捕的载体,利用Red同源重组系统介导的缺口修复技术,分别抓捕牛aSl酪蛋白3’端调控序列(9kb)、hLYZ基因座序列(5kb)、牛axSl酪蛋白5’端调控序列(20kb),使这3个基因片段自动无痕地连接在基因抓捕载体上,形成牛oLSl酪蛋白一hLYZ杂合基因座。结果:实验经过PCR扩增、限制性内切酶酶切验证和序列测定,验证了hLYZ的基因组序列对牛仪S1酪蛋白编码基因组序列的精确置换。结论:这种修复技术为乳腺生物反应器高效表达大载体的制备提供了可行的思路及方法。
简介:目的:与定量比值法比较,探讨全自动直接定量法检测红细胞葡糖-6-磷酸脱氢酶(G-6-PD)活性的可行性。方法:同时采用定量比值法(即硝基四氮唑蓝定量法)和全自动直接定量法,检测219例肝素抗凝静脉血标本的红细胞G-6-PD活性。结果:定量比值法检测G-6-PD缺乏的阳性率为9.13%,全自动直接定量法检测的G-6-PD缺乏阳性率为9.58%,两种方法检测结果无显著性差异(P〉0.05)。结论:定量比值法简单易行,适用于卫生条件有限的基层医疗单位;全自动直接定量法快速准确,是一种可批量检测的理想筛选方法。