1、學(xué)位論文獨創(chuàng)性聲明本人承諾：所呈交的學(xué)位論文是本人在導(dǎo)師指導(dǎo)下所取得的研究成果。論文中除特別加以標注和致謝的地方外，不包含他人和其他機構(gòu)已經(jīng)撰寫或發(fā)表過的研究成果，其他同志的研究成果對本人的啟示和所提供的幫助，均已在論文中做了明確的聲明并表示謝意。學(xué)位論文作者簽名：盞舀薹孥學(xué)位論文版權(quán)的使用授權(quán)書本學(xué)位論文作者完全了解遼寧師范大學(xué)有關(guān)保留、使用學(xué)位論文的規(guī)定，及學(xué)校有權(quán)保留并向國家有關(guān)部門或機構(gòu)送交復(fù)印件或磁盤，允許論文被查閱和借閱。本

2、文授權(quán)遼寧師范大學(xué)，可以將學(xué)位論文的全部或部分內(nèi)容編入有關(guān)數(shù)據(jù)庫并進行檢索，可以采用影印、縮印或掃描等復(fù)制手段保存、匯編學(xué)位論文，并且本人電子文檔的內(nèi)容和紙質(zhì)論文的內(nèi)容相一致。保密的學(xué)位論文在解密后使用本授權(quán)書。學(xué)位論文作者簽名：盔矗，姿指導(dǎo)教師簽名：簽名日期：zal)年廠月70日日本七鰓鰻組織蛋白酶L的克隆表達及功能分析UOmngExpressionandFunctionA11alysisotcathepsinLgene拉Om／一‘‘

3、一‘●一‘^‘‘一^LampetrajaponicaAbstraotCathepsinL(CTSL)belongstothelysosomalcysteineproteasefamily，whichiscomposedofasignalpeptide，apropeptideandamaturepeptideCTSLplaysimportantrolesinavarietyofbiologicalprocesses，suchasprote

4、olysis，antigenpresentingcellapoptosis，TcellsortingaswellasembryonicdevebpmentInadditiontheabnormalchangeoftheexpressionlevelofCTSLaccountsfortheinductionofavarietyoftumors，cardiovascularandkidneydiseaseInparasiticanimals

5、，CTSLmainlyparticipatedinnutritiomlintake，tissuehvasiortimllauneescapeandtissuedifferentiatiorLInthepresentstudy，thefulllengtheDNA(1457bp)ofJapaneselampreyCTSL田CTSL)wasobtainedthroughgenecbning，5and3’endoftherapidamplifi

6、cationexperiments，whichiscomposedbythe5’UTRnoncodingareaof104bp，3UTRnoncodingregionof333bp，andtheopenreadingframeof1020bp，encoding339aminoacidsBioinformaticsanalysisshowedthattheLj—CTSLaminoacidsequencecontainsasignalpep

7、tide(MetlAlal8)，aprecursordomain(Thrj9Glull7)andamaturedomain(AhllSVal339)Thereisapotentialglycosyhtionsites(ASh226)，sixsubstratebindingsites(I_eul86，Met】87，Ah2“，kf80，GIF83andSer335)，andacatalyticsiteconstitutedbythecyst

8、eine(Cysl42)，histidine(His282)andtheasparagine(Ash306)residuesSequencealignmentshowedthattheaminoacidsequenceoftheLjCTSLhasahighhomologywiththeCTSLsfromtheotherspecies，andthehomologywasupto81。1％withtheCTSL舶mXenopuslaevis

9、Besides，巧CTSLcontainsthehighlyconservedmotifs，ERF／WNINandGNFDPhylogenetictreeanalysisshowedthatthe可一CTSLisclosertOtheCTSLfromtheamphibianXenopuslaevis，inaccordancewiththerolesofcreaturesareevolutedfiomlowtOhighHoweverLjC

10、TSLisalsobcatedonitsownbranch，whichdisplaysitsuniqueevolutionarystatusSemiquantitativeRTPCRshowedthatthe碼CTSLiswidelydistributedinthehmpreyheartoralgland，kidneyintestine，livergillsandgonads，andtheexpressionlevelof巧CTSLwa

11、sthemostintheoralgland，whichsuggeststhe巧CTSLplaysimportantrolesinthenutritionalintakeprocessfortheJapaneselampreysThe巧一CTSLwassubcbnedintothepmkaryoticexpressionvectorpTWIN，andsuccessfullyexpressedinEcoliBL21Thesolublere