用光鑷研究微管系統(tǒng)的分子力學(xué)特征.pdf

1、中國農(nóng)業(yè)大學(xué)博士學(xué)位論文用光鑷研究微管系統(tǒng)的分子力學(xué)特征姓名：徐春華申請學(xué)位級別：博士專業(yè)：植物學(xué)指導(dǎo)教師：袁明20050601中國農(nóng)業(yè)大學(xué)博士學(xué)位論文AbstractMicrotubules(MVs)，oneofthemajorcomponentsofnlecytoskeletonineukaryoticcellsplayvitalrolesinmanybiologicalprocessesOneofthemostimportantf

2、unctionsofM1bistogeneratemechanicalforcetopowerVltriOl_lSmovementsofcellsStudyofthemechanicalcharactersofMTbenefitsourunderstandingMTfunctionsandthemechanismofforcegenerationBecauseoftheadvantagesofnoncontactandnonintrus

3、ivemanipulationwithlivingobjects，andmeasuringdisplacementinnanometerscaleandforcein#coNewtonscale，op6caltweezershavebeennowwidelyappliedinbiologicalstudiesInthisdissertation，adualopticaltweezerssystemwasusedtostudythemol

4、ecularmechanicalcharactersofmicrotubulesystem，includingthebreakingforceoffuorescentlabeledmicrotubule，theprocessofthephotosensitivebreakage，theelasticityofMTthemechanismsofthephotosensitivebreakageandtheunbindingforcebet

5、weenmicrotubuleassociateproteins(MAPs)andMrsThebiologicalsystempracticableforopticaltweezersstudieswasfirstsetupTubulinWaspurifiedfromporcinebrainandlabeledbyrhodamineandbiotinMTswereassembledinvitroandstabilizedbytax01D

6、uringexperiments，wefoundfuorescancelabeledMrsbrokeunderexcitationlightTesolvethisproblem，weinvestigatedthemechanismofhd丌cleavageindetailTheresultsshowedthatfluorescentdyeandexcitationlightwerebethnecessarytothephotosensi

7、tivebreakageThestrongerthelightintensityandthehigherthedyeconcentration，thefasterbrokeuptheNITsItisthereactiveoxygenspecies(ROS)producedbytheinteractionbetweenthedyesandtheirexcitationlightledtotheMTbreakageFreeradicalso

8、nekindofROS，werethemaindestroyersAscorbicacidrestrainedthefluorescentMTbreakageeffectivelyTheoperationsystemWaSthensetupincludingthesamplecellsandmicmsphereforthemanipulationVarioussamplecellswelemadeaccordingtodifferent

9、experimentsFlowcellandtemperaturecontrolflowcellwerealsomadeforreplacementofthebufferandcontrollingtemperatureOpticaltweezers‘caught’singleNITthroughmicroshpereswhichWerecoatedwithstrepavidintobindtotheMTAndthemechanical

10、forceWasmeasuredbytheopticaltweezersFluorescencelabeledMTswereelongatedabout20％抽averagewhenstretchedbyopticaltweezersWhensomeprotofilamentsoftheMTbrokeuptheresidualprotofilamentswasstretchedevenlongerThisresultindicatedt

11、heelasticpropertyoffluorescencelabeledMltThepersistencelengthoffluorescencelabeledMTwas3O“l(fā)ersofmagnitudeshorterthanthatofMrswithoutnuoRsc釘lcelabeled，ItWaSaffectedbytaxolandexcitationlightThebreakingforceobtainedbytheopt