簡(jiǎn)介：東北農(nóng)業(yè)大學(xué)碩士學(xué)位論文中藥透皮劑對(duì)實(shí)驗(yàn)性和臨床型乳腺炎治療效果研究姓名杜瓊申請(qǐng)學(xué)位級(jí)別碩士專業(yè)基礎(chǔ)獸醫(yī)學(xué)指導(dǎo)教師張秀英20080620ABSTRACTSTUDIESONTHERAPEUTICEFFICACYOFTRADITIONALCHINESEMEDICINEPERCUTANEOUSONMOUSETENTATIVEMASTITISANDCOWCLINICALMASTITISABSTRACTDAIRYCOWMASTITISISONEOFTHEMOSTCOMMONDISEASES；NCOWCULTIVATION．ITNOTONLYINFLUENCESTHEQUANTITYOFCOWMILKPRODUCTION，CAUSESSEVEREECONOMICLOSSES，BUTALSOINFLUENCESTHEQUALITYOFTHEMILK．PEOPLEHAVEPAYEDCLOSEATTENTIONTOTHEPREVENTIONOFTHECOWMASTITIS．ATPRESENTTHECOWMAINPATHOGENICBACTERIAOFMASTITISARESTAPHYLOCOCCI，STREPTOCOCCIANDESCHERICHIACOLI．USUALLYTHEANTIBIOTICSWEREUSEDTOCURETHECOWMASTITIS，HOWEVER,THEPROBLEMSOFRESIDUESANDBACTERIARESISTANCETOANTIBIOTICSHAVEBECOMINGMOREANDMORESERIOUSWHENANTIBIOTICWEREUSED，WHICHHAVEBEENWORRIEDBYTHEPEOPLEOFALLWORLD．CHINESEMEDICINALHERBDOESNOTYIELDTOLERANCEANDHAVETHENATUREOFLOWPOISONANDANTIINFLAMMATORYANDSOON．THETHERAPYOFCOWMASTITISWITHCHINESEMEDICINALHERBISFOLLOWEDINTERESTBYMANYSCHOLARS．TRADITIONALCHINESEMEDICINEPERCUTANEOUSISANEXTERNALPREPARATIONADVANCEDDEVELOPMENTBYOURLABORATORY，ITMAINCONTAINSDANDELION、FORSYTHIASUSPENSAANDBAICALSKULLCAPROOT，HASHEAT。CLEARINGANDDETOXICATING，ANTIBIOSISANDANTIPHLOGISTIC，DETUMESCENCEANDACESODYNEFUNCTION．INORDERTOESTIMATETHETHERAPEUTICEFFICACYOFTHISPREPARATIONONTHEMASTIFFS，WECONDUCTEDTHISTOPICRESEARCH．55BACTERIALSTRAINSWEREISOLATEDFROM41MILKSAMPLESOFCLINICALMASTITIS，INCLUDINGSTAPHYLOCOCCI20OF55，36．4％，STREPTOCOCCI24OF55，43．6％ANDGRAM．NEGATIVEBACILLI11OF55，20喲．THELEADINGPATHOGENSOFCLINICALMASTITISWERESTREPTOCOCCUSAGALACTIAE，STAPHYLOCOCCUSAUREUSANDESCHERICHIACOLIACCOUNTINGFOR30．9％．21．8％AND16．3％．BYUSINGTHEAPPROACHOFARTIFICIALCHALLENGEMOUSE，0．05ML106CFUML。1BACTERIUMLIQUIDWHICHCONTAINTHREEKINDSBACTERIUMWEREINJECTEDINTOMAMMARYGLAND．24HOURSLATER，MOUSEEMERGEDAPPARENTCLINICALSYMPTOMANDPATHOLOGICALCHANGE。THECONTENTOFTNFNANDACTIVITYOFNAGASEINMAMMARYGLANDTISSUERISEDOBVIOUSLY,ANDBACTERIUMSWEREISOLATEDFROMTHEMAMMARYGLANDWHICHWEINJECTEDBACTERIUMS．ITDEMONSTRATEDTHATWESUCCEEDMAKINGMOUSETENTATIVEMASTITIS．THROUGHTHETHERAPEUTICTESTONMOUSETENTATIVEMASTITIS，THETHERAPEUTICEFFICACYOFTRADITIONALCHINESEMEDICINEPERCUTANEOUSWASESTIMATED．THERESULTSSHOWEDTHECURERATEANDEFFECTIVEPOWEROFTRADITIONALCHINESEMEDICINEPERCUTANEOUSWERE60％AND80％，ITCOULDOBVIOUSLYLLL

簡(jiǎn)介：東北農(nóng)業(yè)大學(xué)博士學(xué)位論文奶山羊乳腺發(fā)育的形態(tài)學(xué)研究姓名曲波申請(qǐng)學(xué)位級(jí)別博士專業(yè)動(dòng)物生物化學(xué)與分子生物學(xué)指導(dǎo)教師李慶章20080620東北農(nóng)業(yè)大學(xué)農(nóng)學(xué)博上學(xué)位論文到妊娠期迅速升高，整個(gè)泌乳期B酪蛋白表達(dá)水平比較平穩(wěn)，各時(shí)點(diǎn)無(wú)差異，而其基因表達(dá)水平卻有顯著差異，可見(jiàn)乳腺中P酪蛋白的表達(dá)調(diào)控可能涉及多種調(diào)節(jié)岡素的參與退化早期B酪蛋白及其基因表達(dá)水平開(kāi)始逐漸下降，但仍顯著高于青春期，可能是為下一次妊娠做準(zhǔn)備，更快進(jìn)入分泌表達(dá)狀態(tài)。此外，本研究采用活細(xì)胞熒光標(biāo)記法結(jié)合激光共聚焦顯微技術(shù)，在Ⅱ細(xì)胞水平觀察奶山羊乳腺發(fā)育中的結(jié)構(gòu)變化。結(jié)果顯示奶山羊乳腺發(fā)育中，腺上皮細(xì)胞中內(nèi)質(zhì)網(wǎng)和線粒體的變化規(guī)律與乳腺組織超微結(jié)構(gòu)和乳成分組化檢測(cè)結(jié)果基本相符。退化期乳腺上皮細(xì)胞內(nèi)仍維持一定數(shù)量的細(xì)胞器，可能是維持細(xì)胞正常機(jī)能和活動(dòng)所必需的。關(guān)鍵詞奶山羊；乳腺發(fā)育；P一酪蛋白；激光共聚焦顯微技術(shù)

簡(jiǎn)介：鄭州大學(xué)博士學(xué)位論文TPA乳腺特異性表達(dá)載體的構(gòu)建及其瞬時(shí)表達(dá)姓名丁一申請(qǐng)學(xué)位級(jí)別博士專業(yè)病理與病理生理學(xué)指導(dǎo)教師張欽憲20040428鄭州人學(xué)2004脯博I例冗生論文TPA乳腺特異性衰叢戴怍的構(gòu)建披』L瞬時(shí)表達(dá)乳腺生物反應(yīng)器MAMMARYGLANDBIOREACTOR技術(shù)是利用哺乳動(dòng)物乳腺特異性表達(dá)的啟動(dòng)子元件構(gòu)建轉(zhuǎn)基兇動(dòng)物，指導(dǎo)外源基因在乳腺中表達(dá)，并從轉(zhuǎn)基因動(dòng)物的奶液中獲取重組蛋白。利用乳腺生物反應(yīng)器生產(chǎn)藥用蛋白具有表達(dá)體系簡(jiǎn)前、町進(jìn)行翻譯后修飾及費(fèi)用低廉等優(yōu)點(diǎn)。乳腺生物反應(yīng)器己成為生產(chǎn)重組蛋白的首選手段。建立乳腺生物反應(yīng)器的關(guān)鍵在于構(gòu)建乳腺特異性表達(dá)載體。乳腺特異性表達(dá)載體由乳腺特異性啟動(dòng)子、目的基因和加POLYA信號(hào)三部分紕成。目前已克隆并用作構(gòu)建載體的乳蛋白基因主要包括B一乳球蛋白基因BLG、酪蛋白基因和乳清酸蛋白基因WAP等。本研究中，首先將克隆的乳清酸蛋白基岡WAP24KB啟動(dòng)子插入PBLUESCRIPT質(zhì)粒，構(gòu)建質(zhì)粒PW；然后將克降的037KB加POLYA信號(hào)插入WAP啟動(dòng)子的下游，構(gòu)建I_LR指導(dǎo)外源基岡在乳腺特異性表達(dá)的載體PWA；最后將克隆的目的基因21KBTPACDP4A連接于UP啟動(dòng)子和加POLYA信號(hào)之間，構(gòu)建PA乳腺特異性表達(dá)載體PWTA。構(gòu)建成功的PWTA表達(dá)載體通過(guò)脂質(zhì)轉(zhuǎn)染胺試劑LIPOFECTAMINE。”“OO轉(zhuǎn)染乳腺癌細(xì)胞系，通過(guò)體外注射轉(zhuǎn)移至妊娠期小鼠乳腺，以原位雜交技術(shù)檢測(cè)TPA在乳腺癌細(xì)胞系和泌乳期小鼠乳腺中的瞬時(shí)表達(dá)，驗(yàn)證PWTA表達(dá)載體的表達(dá)能力。材料和方法1載體PW的構(gòu)建以含WAP啟動(dòng)子的質(zhì)粒WAP2H；H為模板，設(shè)計(jì)引物上、下游引物分別帶有NHEI和ITINDLII酶切位點(diǎn)，PCR擴(kuò)增WAP啟動(dòng)子DNA片段；以NHEL和HJNDLLL對(duì)擴(kuò)增產(chǎn)物進(jìn)行雙酶切，瓊脂糖凝膠電泳回收2，4KB酶切片段。以XBAI和HINDLII雙酶切PBLUESCRIPT質(zhì)粒，瓊脂糖凝膠電泳回收297KB酶切片段。NHEI和XBAT粘往末端互補(bǔ)。以體外連接試劑盒連接24KBWAP肩動(dòng)子片段和297KBPBLTLESCRPL質(zhì)粒DNA片段，16。C，連接過(guò)夜。重組體轉(zhuǎn)化感受念JML09細(xì)菌。將轉(zhuǎn)化菌涂于含AMP的培養(yǎng)皿，37“C，培養(yǎng)過(guò)夜。隨機(jī)挑選10個(gè)克隆，接種于含AMP的LB培養(yǎng)基，37℃，培養(yǎng)過(guò)夜。小提質(zhì)粒，分別進(jìn)行TTINDIII單酶切和HINDIIL／SAC11雙酶切鑒定。構(gòu)建成功的載體命名為PW。2載體PWA的構(gòu)建以PCDNA30質(zhì)粒為模板，設(shè)計(jì)引物T、卜游引物

簡(jiǎn)介：軍事醫(yī)學(xué)科學(xué)院碩士學(xué)位論文牛ΑS1酪蛋白基因序列指導(dǎo)的人組織型纖溶酶原激活劑突變體微小基因在小鼠乳腺中的表達(dá)姓名譚曉紅申請(qǐng)學(xué)位級(jí)別碩士專業(yè)細(xì)胞生物學(xué)指導(dǎo)教師鄧?yán)^先200061BOVINEASLCASEINGENESEQUENCESDIRECTEXPRESSIONOFAVARIANTOFHUMANTISSUEPLASMINOGENACTIVATORINTHEMILKOFTRANSGENICMICEABSTRACTTHEBIRTHOF“SUPERMICE”BROKETHEVALLATIONOFSPECIESINBIOSPHERE，ANDTHERESEARCHOFTRANSGENICANIMALACHIEVEDDRAMATICPROGRESSTRANSGENICPHARMACYSPDNGUPFOLLOWINGTHESTUDIESOFTHEMAMMARYGLANDBIOREACTORTHEMAMMARYGLANDOFTRANSGENICLIVESTOCKISLIKELYTOSERVEASTHEBIOREACTORFORPRODUCINGTHEBIOACTIVEPOLYPEPTIDEMEDICINEORPROTEINWITHSPECIALNUTRITIONINTLLISSTUDYTHEPROMOTEROFCASEINGENEWASUSEDTODIRECTTHETPAMINIGENETOEXPRESSINTHEMAMMARYGLANDSPECIFICALLYTHEMBTANTTPAWASDETECTEDINTHEMILKOFTHETRANSGENICMICE111ESEQUENCESOF58KBCOMPRISINGOFTHESEQUENCESFROMEXORT2TOEXON6OFTPAWEREAMPLIFIEDFROMHUMANGENOMICDNABYPCRLATPAMINIGENEWASGENERATEDBYLIGATINGTHEGENOMICSEQUENCESOFTPATOTHELATPACDNAATTHESITEOFNARIALPHASLCASEINVECTORWASMODIFIEDBYDELETINGTHESEQUENCESFROMTHESTARTINGSIGNALCODEOFTRANSLATIONANDAFTERITANXHOISITEWASINTRODUCEDATTHE3’ENDOFCASEINGENEOFTHEVECTORTOFACILITATETHESEPARATIONOFFUSIONGENEFROMTHEPROKARYOTICSEQUENCESFORMIRCOINJECTIONNLEMAMMARYGLANDSPECIFICEXPRESSIONCONSTRUCTWASOBTAINEDBYINSERTINGTHELATPAMINIGENEINTHEMODIFIEDCASEINVECTORNLEFUSIONGENECONTAININGTHEPROMOTEROFCASEINGENELATPAMINIGENEANDTHEDOWNSTREAMOFCASEINGENEWASINTRODUCEDINTOTHEFERTILIZEDEGGSOFMICEBYMICROINJECTIONTHEINJECTEDEGGSWEREFURTHERTRANSPLANTEDINTOTHEOVIDUCTSOFRECIPIENTMICEABONT2700FERTILIZEDEGGSWEREINJECTED，AND2200EGGSWERETRANSPLANTEDINTO81RECIPIENTMICEOF47OFFSPRING，F(xiàn)IVEWEREPOSITIVETRANSGENICMICEN心CONCENTRATIONOFLATPAINTHEMILKOFONEFEMALETRANSGENICMOUSEIS018UG／M1THISRESULTSHOWEDTHAT也ELATPAMINIGENECOULDCOITCCUYEXPRESSTHEBIOACTIVELATPAINTHEMILKOFTHETRANSGENICMOUSEUNDERTHECONTROLOFCASEINGENE2

簡(jiǎn)介：軍事醫(yī)學(xué)科學(xué)院博士學(xué)位論文牛Β乳球蛋白基因調(diào)控成分的克隆和用以制備小鼠乳腺生物反應(yīng)器的研究姓名陳紅星申請(qǐng)學(xué)位級(jí)別博士專業(yè)遺傳學(xué)指導(dǎo)教師蘇國(guó)富200061達(dá)到12UG／ML，最低則無(wú)法檢測(cè)到TPA活性。這個(gè)結(jié)果顯示位點(diǎn)效應(yīng)對(duì)TPA的表達(dá)具有重大的影響，也提示表達(dá)載體中用以對(duì)抗位點(diǎn)效應(yīng)的MAR序列并未起作用。上述的結(jié)果表明／本文克隆的牛B乳球蛋白基因表達(dá)調(diào)控序列可以控制目的基因在轉(zhuǎn)基因小鼠中表達(dá)，并且產(chǎn)物能分泌到乳汁中，可以用以制備乳腺生物反應(yīng)器。關(guān)鍵詞生物反應(yīng)器TPAB乳球蛋白2

簡(jiǎn)介：中國(guó)農(nóng)業(yè)大學(xué)博士學(xué)位論文擬蜘蛛牽絲蛋白基因在大腸桿菌和小鼠乳腺表達(dá)研究姓名許紅韜申請(qǐng)學(xué)位級(jí)別博士專業(yè)生物化學(xué)與分子生物學(xué)指導(dǎo)教師李寧20040601ABSTRACTSPIDERDRAGLINESILKISALLIMPRESSIVEMATERIALWITHACOMBINATIONOFTENSILESTRENGTHANDELASTICITYTHATENDOWSITWITHUNIQUETOUGHNESSANDHIGHERENERGYTOBREAKTHANANYOTHERCOMMONMATERIAL，NATURALORARTIFICIALDRAGLINESILKISFIVETIMESSTRONGERBYWEIGHTANDLIGHTERTHANSTEELBECAUSEOFITSEXCELLENTMECHANICALPROPERTIESTHEDRAGLINESILKCOULDBEUSEFULFORINDUSTRIAL，MILITARYMATERIAL，WEAVEANDMEDICALPURPOSESTHISRESEARCHFOCUSESONACQUIRINGARTIFICIALSPIDERDRAGLINESILKPROTEINGENEWHICHCANEXPRESSTARGETPROTEINEFFICIENTLYANDCALLBEPERFORMEDEASILY，ANDTHENPRODUCESPIDERSILKINLARGEQUANTITIESFORINDUSTRIALUSEACCORDINGTOTHEKNOWNPARTIALCDNASEQUENCEOFDRAGLINESILKTWOARTIFICIALDRAGLINESILKGENEMONOMERS，A360BPSEQUENCEANDA390BPSEQUENCE，WEREDESIGNEDTOENCODEANALOGSOFTHEPROTEINOFNEPHILACLAVIPESDRAGLINESILKDNAMONOMERSEQUENCESWEREMULFIMERIZEDTOENCODEHIGHMOLECULARWEIGHTARTITICIAISPIDERDRAGLINESILKPROTEINTWOTIMERFOTTRTIMERSIXTIMEREIGHTTIMERANDSIXTEENTIMERWERERESPECTIVELYGAINEDACCORDINGTOTILEKNOWNPARTIALAMINOACIDSEQUENCEOFDRAGLINESILK，ANAN『LINOACIDSEQUENCEOF19AAWASSYNTHESIZEDANTISERUMAGAINSTDRAGLINESILKPROTEINWASRAISEDINRABBITSAGAINSTS3RNTHETIEPEPTIDESCONJUGATEDTOKEYHOLELIMPETHEMOCYANINEIGHTTIMERANDSIXTEENTIMEROFTWOARTIFICIALDRAGLINESILKGANEMONOMERSWERECLONEDINTOPET30AVECTORAPROKARYOTICEXPRESSIONVECTORANDINDUCEDWITHIPTGFOUREXPRESSIONVECTORPET30AX8、PET30AX16、PET30AF8ANDPET30AF16WERECONSTRUCTEDEACHSTRAINPRODUCEDACHARACTERISTICHETEROGENEOUSARRAYOFIMNLUNOREACTIVEBANDSTHEMASSESOFTHESTRONGESTBANDWERERESPECTIVELY85KD、165KD、85KDAND165KD，WHICHREPRESENTSTHEFULLLENGTHGENEPRODUCTINEACHPATCERNTHECONCENTRATIONSOFRECOMBINANTDRAGLINESILKPROTEINWERERESPECTIVELY800MGL、500MG／L、200MG／LAND200MG／LACCORDINGTOGOAT，EASEINSIGNALSEQUENCE，A125BPDNASEQUENCEWASDESIGNEDANDSYNTHESIZEDFOURTIMERANDSIXTIMEROFTWOARTIFICIALDRAGLINESILKGENEMONOMERSWERELIGATEDWITHTHESIGNALSEQUENCEANDTHENWERECLONEDINTOPBCLVECTOR，WHICHISASPECIFICMILKEXPRESSIONVECTORFOUREXPRESSIONVECTORPBCIF4，PBCIX4PBCLF6ANDPBCIX6WERECONSTRUCTEDTRANSGENIEMICEWEREGENERATEDBYMICROINJEETIONOFFERTILIZEDOOCYTEWITHTWOCONSTRUCTS，PBCIF4ANDPBCLX6FORPBCIX6VECTORTRANSGENICMICEWERESCREENEDBYPERANDSOUTHERNBLOTWHICHREVEALEDTHAT10MICE5毋5早AMONG繇MICEWERE打ANSGENICPOSITIVETHEINTEGRATIONRATEIS17％FIMICEFORMOSTOFFOUNDERLINEWEREALSODETECTEDBVPCRANDTHEPOSITIVETRANSGENICMICEWEREFOUNDMILKOFFIVEF0MICEANDEIGHTF1MICEWASANALYZEDBYWESTERNBLOTANDRADIOIMNMOASSAYTWOF0MICEANDSEVENFLMICEEXPRESSEDRECOMBINANTDRAGLINESILKPROTEINTHEMOLECULARNLASSESOFEXPRESSEDRECOMBINANTDRAGLINESILKPROTEINBYMOSTOFMICEWEREABOUT55KD，WHICHWASCONSISTENTWITHTHATOFTHEORETICCALCULATIONBUTSOMEMICEEXPRESSEDRECOMBINANTDRAGLINESILKPROTEINWHICHMOLECULARMASSESWERENOTCONSISTENTWITHTHATIⅡ

簡(jiǎn)介：鄭州大學(xué)碩士學(xué)位論文Β胡蘿卜素誘導(dǎo)人乳腺癌MCF7細(xì)胞凋亡與PPARΓ信號(hào)傳輸及活性氧產(chǎn)生的聯(lián)系姓名崔艷紅申請(qǐng)學(xué)位級(jí)別碩士專業(yè)應(yīng)用化學(xué)指導(dǎo)教師趙文恩20060415使用50岍10L幾P一胡蘿卜素，當(dāng)將處理時(shí)間從L天延長(zhǎng)至Ⅱ4天時(shí)，細(xì)胞活力從71％降為42％。作為PPAR吖抑制劑的GW9662和／或GSH能夠削弱P．胡蘿H素誘導(dǎo)的細(xì)胞活力的下降，5HM01／LGW9662預(yù)處理能部分挽回P一胡蘿B素誘導(dǎo)的細(xì)胞活力的下降，而10肛M01幾GW9662預(yù)處理部分增強(qiáng)B．胡蘿H素誘導(dǎo)細(xì)胞活力的下降。GSH能呈劑量效應(yīng)抑制P一胡蘿H素對(duì)細(xì)胞活力的抑制作用，用200岬OL，LGSH預(yù)處理細(xì)胞，能使細(xì)胞活力由50％恢復(fù)到70％，且5岬OL／LGW9662共同預(yù)處理能增加GSH對(duì)細(xì)胞的保護(hù)作用。50岫OL幾P．胡蘿H素處理3天能顯著增加細(xì)胞早期的凋亡率，用GW9662或GSH能削弱這種作用，且共同使用GW9662和GSH有協(xié)同作用。B一胡蘿H素對(duì)MCF一7細(xì)胞的周期沒(méi)有明顯的影響。4．B胡蘿H素誘導(dǎo)MCF．7細(xì)胞的細(xì)胞核形態(tài)的變化P一胡蘿H索處理的細(xì)胞，細(xì)胞核變小，濃縮，熒光加強(qiáng)和成核碎片等，細(xì)胞呈凋亡狀態(tài)。200¨MOL幾GSH預(yù)處理能明顯減少細(xì)胞核的形態(tài)變化，200啪。此GSH和5岬OL／LGW9662預(yù)處理能明顯減少細(xì)胞核的形態(tài)變化。單獨(dú)用GW9662預(yù)處理，對(duì)由P胡蘿N素誘導(dǎo)的細(xì)胞核的變化的影響不明顯。5．B一胡蘿B素使細(xì)胞內(nèi)活性氧水平上升用50岬OI／LP一胡蘿H素處理3天后，細(xì)胞內(nèi)活性氧水平明顯比對(duì)照組高，且用200肚M01幾GSH預(yù)處理能顯著抑制這種升高。用GW9662預(yù)處理不能降低由B．胡蘿H索引起的活性氧的增加，并且LO“MOL／LG、Ⅳ9662能使細(xì)胞內(nèi)活性氧增加為對(duì)照組的120％。、6．B一胡蘿B素誘導(dǎo)細(xì)胞色素C的釋放50肛MOL／LP一胡蘿卜素處理3天后，紅色和綠色有所分離，說(shuō)明細(xì)胞色素C從線粒體內(nèi)釋放出來(lái)，跑到了胞漿里。通過(guò)WCSTEMBLOT檢測(cè)，進(jìn)一步表明B．胡蘿H素誘導(dǎo)細(xì)胞色素C的釋放，200I_IMOI／LGSH或200肛MOL／LGSH5岬01／LGW9662能夠抑制細(xì)胞色素C的釋放。綜合前人的工作及我們的實(shí)驗(yàn)數(shù)據(jù)，我們有理由認(rèn)為，PPAR．T傳輸系統(tǒng)和活性氧的產(chǎn)生，可能在P胡蘿卜素抑制細(xì)胞生長(zhǎng)和誘導(dǎo)凋亡方面的分子機(jī)制中都起到了重要的作用。并且，由于GW9662和GSH共同預(yù)處理作用效果更好，這兩個(gè)途徑可能有協(xié)同作用。II

簡(jiǎn)介：Y977729分類號(hào)Q21UDC52Z密級(jí)壘玨級(jí)⑧西北農(nóng)林針捉大學(xué)2006屆攻讀碩士學(xué)位研究生學(xué)位畢業(yè)論文人溶菌酶基因乳腺特異性表達(dá)載體的構(gòu)建及其在小鼠乳腺癌細(xì)胞中的表達(dá)學(xué)科專業(yè)筮直生物堂研究方向喧乳動(dòng)麴艇臉王程研究生值塵寶指導(dǎo)教師韭涵麴援完成時(shí)間2壁豎生查且CONSTRUCTION0FMA～口MARYGLANDSPECIFICEXPRESS10NALVECTORANDEXPRESS10NINMOUSEBREASTCARCINOMACELLSFORHUMANLYSOZYMEABSTRACTHUMANLYSOZYMEHLYZPARTICIPATEINTHEBODYDEFENSEMECHANISMITACTSASASHIELDAGAINSTBACTERIALINFECTIONANDITHASEFFECTSONMALIGNANTGROWTHANDIMMUNOLOREGULATIONSOITHASWIDEAPPLICATIONINMEDICINEANDFOODSTUFFINDUSTRYHLYZCANBEPURIFIEDFROMHUMANBREASTMILK，NEUTROPHILS，ANDURINEBECAUSEOFTHELIMITATIONOFITSSOURCEANDTHECOMPLEXITYOFITSEXTRACTION，ANDTHEPOTENTIALRISKOFTRANSMITTINGPATHOGENSTOTHENURSINGINFANTS，WECANNOTGETITBYLARGESCALESODEVELOPMENTOFSAFERRECOMBINANTHLYZISNEEDEDANIMALMAMMARYGLANDBIOREACTORISANOVELALTERNATIVESTRATEGYFORPRODUCTIONOFRECOMBINANTPROTEINSWITHMANYADVANTAGESINCLUDINGHIGHPRODUCTIONCAPACITIES，EASYPURIFICATION，F(xiàn)AITHFULTRANSLATIONALMODIFICATIONSANDAUTHENTICBIOLOGICALACTIVITIESOFTHEEXPRESSEDPRODUCTSINTHISRESEARCH，THEMAMMARYGLANDSPECIFICEXPRESSIONVECTORPEBHWASCONSTRUCTED，WHICHWASCONSISTEDOFREGULATINGELEMENTSREGULATIONREGIONOFBOVINE3CASEINANDEXPRESSIONSEQUENCEHUMANLYSOZYMECDNATHENPEBHWASTRANSFECTEDINTOMOUSEBREASTCARCINOMACELLSCULTURED加VITROANDTHEEXPRESSIONCELLSTRAINSWEREABTAINEDTHEEXPRESSEDPRODUCTSWEREANALYSEDBYSDSPAGEANDWESTERNBLOTTHERESULTSOFRESEARCHAREASFOLLOWING1HUMANLYSOZYMECDNAWASCLONEDFROMHUMANPLACENTATOTLERNAWITHRTPCR，WHICHWAS853BPANDCOMPRISEDINTACTCODINGREGIONSSEQUENCEANALYSISDEMONSTRATEDTHATITSHOMOLOGY、VITHTHERELATIVEREGIONOFHUMANLYSOZYMEONGENBANKWAS99％WITHINTHEOPENREADINGFRAME，THREESITEMUTATIONSOCCUREDWHICHCOULDNOTALTERPROTEIN’SFUNCTION2BYTHESTRATEGYOFDIRECTIONALCLONINGHLYZWASFUSEDWITHTHECOMMONLYPRIMARYEXPRESSIONVECTORPBCPTHENWASCLONEDDIRECTIONALLYINTOEXPRESSIONVECTORPEGFPCI，THEMAMMARYGLANDSPECIFICEXPRESSIONVECTORPEBHFORHLYZWASCONSTRUCTED，WHICHINCLUDEDRESISTANTGENEANDEGFPREPORTERGENEITCOULDALSOENSUREEGFPANDAIMEDGENEEXPRESSRESPECTIVELY3THEVECTORPEBHWASTRANSFECTEDINTOMOUSEBREASTCARCINOMACELLSCULTUREDINVITRO

簡(jiǎn)介：山東大學(xué)碩士學(xué)位論文苯達(dá)莫司汀（BENDAMUSTINE）誘導(dǎo)小鼠乳腺癌細(xì)胞凋亡及其機(jī)制的研究姓名孔芳申請(qǐng)學(xué)位級(jí)別碩士專業(yè)細(xì)胞生物學(xué)指導(dǎo)教師孔慶忠20070506山東大學(xué)碩士學(xué)位論文中文摘要苯達(dá)莫司汀BENDAMUSTINE誘導(dǎo)小鼠乳腺癌細(xì)胞凋亡及其機(jī)制的研究背景乳腺癌是人類最常見(jiàn)的惡性腫瘤之一，其發(fā)病率近年來(lái)呈上升趨勢(shì)。在多種治療乳腺癌的方法中，化療仍是最為常用且效果較明顯的重要手段之一然而現(xiàn)有的化療藥物或多或少都會(huì)引起藥物耐受性的產(chǎn)生，因此，尋找有效殺傷乳腺癌細(xì)胞的化療藥物已成為研究者當(dāng)前的主要任務(wù)之一臨床研究表明，作為氮芥類藥物且具多種功能的烷化劑，苯達(dá)莫司汀BENDAMUSTINE對(duì)乳腺癌有較好的療效然而，相關(guān)的研究數(shù)據(jù)有限，作用機(jī)制不清楚由于很多小分子化療藥物可通過(guò)影響腫瘤細(xì)胞周期和誘導(dǎo)細(xì)胞凋亡來(lái)抑制腫瘤細(xì)胞的生長(zhǎng)，因此我們重點(diǎn)研究苯達(dá)莫司汀對(duì)體外培養(yǎng)的小鼠乳腺癌C127細(xì)胞周期和誘導(dǎo)凋亡的作用及作用機(jī)制，以期為腫瘤生物學(xué)和腫瘤化療提供理論依據(jù)目的觀察抗癌藥物苯達(dá)莫司汀對(duì)體外培養(yǎng)的小鼠乳腺瘤細(xì)胞系D127生長(zhǎng)抑制及其作用機(jī)制，以期在細(xì)胞水平上為治療乳腺癌提供相關(guān)的理論依據(jù)方法；細(xì)胞增殖檢測(cè)方法采用形態(tài)學(xué)觀察和細(xì)胞記數(shù)法檢測(cè)苯達(dá)莫司汀對(duì)C127細(xì)胞形態(tài)和存活率的影響；細(xì)胞凋亡檢測(cè)方法用吖啶橙染色法，在熒光顯微鏡下觀察細(xì)胞核的形態(tài)變化和凋亡小體；TUNEL法檢測(cè)DNA的片段化；細(xì)胞凋亡和壞死的鑒別檢測(cè)乳酸脫氫酶CIDVI的釋放區(qū)分壞死和凋亡；P53蛋白表達(dá)測(cè)定免疫熒光結(jié)合激光共聚焦掃描顯微技術(shù)測(cè)定細(xì)胞中P53的表達(dá)情況；細(xì)胞周期的變化流式細(xì)胞術(shù)檢測(cè)苯達(dá)莫司汀對(duì)C127細(xì)胞周期的影響結(jié)果1相差顯微鏡觀察顯示苯達(dá)莫司汀20／ⅡG／ML作用C127細(xì)胞24H之后細(xì)胞形態(tài)無(wú)明顯變化，48H后細(xì)胞表現(xiàn)出凋亡的特征，細(xì)胞突起變少，細(xì)胞中顆粒和3

簡(jiǎn)介：新疆農(nóng)業(yè)大學(xué)碩士學(xué)位論文位點(diǎn)不依賴性乳腺表達(dá)載體的構(gòu)建及在乳腺上皮細(xì)胞中的表達(dá)姓名胡艷秋申請(qǐng)學(xué)位級(jí)別碩士專業(yè)生化與分子生物學(xué)指導(dǎo)教師羅淑萍李文蓉20060601CONSTLLLCTIONANDCEUEXPRESSI蛐OFMAMMARYGLANDEXPRESSIONVBCTORWITHPOSITIONINDEPENDENTMANNERHUYANQIUABSTRACTTHEPURPOSEOFTHISPAPERWASTOCONSTMCTMAMMARYGLANDEXPRESSIONVECTORWITHPOSITIONINDEPENDENTMALLILER，WHICHCANPMBCTTRANSGENESFROMGENOMICPOSJFIONEFI色CTSANDEXPRESSINDEPENDENTLY．THEENDOGENOUS0一CASEINHADBEENAⅡALYZEDBYR1二PCRREVERSETRANSCRIPTIONPOIYMERASECHAINREACTIONFORTHEASSESSMENROFTHEFLLNCTIONOFMAMMARY印ITHELIALCELLSMECANDTHEEFFICACYOFHO蛐ONALINDUCTIONINVITMOPTIMIZEDVECTORPWHERE2V．01WASRECONSTMCTEDBYEXDSINGBCZGENEFIRSTLYANDTHEREAFTERTHEBLGVP2GENEWASINSERTEDIN枷LTIPLECLONESITEMCSFLANKINGBYINSULATORS．THUSAMAMMARYGLANDEXPRESSIONVECTORTHATTRIEDTOPROTECTTRANSGENESFROMGENOMICPOSITIONEFFECTSWASCONSTNLCTED．GOATM鋤MARYEPITLLELIALCELLSGMECWEREISOLATEDFROMLACTATINGGOATMAMMARYTISSUE．RT_PCRANDIMMUNOCHEMICALMETHODWEREUSEDTODETECTBCASEINEXPRESSION．THETRANSCRIPTIOⅡOFTWOMAMMARYEPITHELIALCEULINESC127ANDT47DWASALSOANALYZEDTOCOMPAREANDASSESSTHEPCASEINGENEEXPRESSIONINDI丘EFENTMAMMARY印ITHELIALCELLLINES．THERESULTSSHOWEDTHATPCASEINGEⅡEEXPRESSIONCANBEDETECTEDINPRIMARYCULTUREDGOATMAMMARYEPITHELIALCELLSGMEC1DERIVEDFROMLACTATING90ATASWELLASCONSEQUENNYPASSAGEDCELLSGMEC一3WITHTHEPRESENCEOFHORINOILESINCLUDINGPROLACTIN，INSULINANDHYDROCORTISONE2

簡(jiǎn)介：西北農(nóng)林科技大學(xué)碩士學(xué)位論文STAT3基因轉(zhuǎn)染牛乳腺上皮細(xì)胞及轉(zhuǎn)染后細(xì)胞的生物學(xué)特性觀察姓名劉甲申請(qǐng)學(xué)位級(jí)別碩士專業(yè)發(fā)育生物學(xué)指導(dǎo)教師張涌20080501THEB10LOGICALCHARACTERISTICSOFBOVN呵EMAMMARYEPITHELIALCELLSTRANSFECTEDWITHSTAT3GENEABSTRACTSTATSARELATENTTRANSCRIPTIONFACTORS也ATMEDIATECUT拋NE鋤DGROWTHFACTORDIRETED仃蜀11LS謝PTIONKMANYHMNALLCANCERS吼D仃觚SFOMED∞LLLINES，STAT3ISPERSISTENTLYACTICATED，ANDINCENCULTⅧC，ACTIVESTAT3ISEITHERREQUIRED南R仃ANSFOMATION，E11HANCES仃孤SF0MATION，ORBLOCKSAPOPTOSISSTAT3GELLESWERE仃ALLSCRIPTIONFACTORSRALHERACTIVCLYSTLLDIEDINRECEILTYEARSMANYRESE∞CHRESULTSSHOWEDTHATSTAT3EXPRESSEDABNONNALLYINLOTSOF劬MORTISSUESANDCEUSYSTEMS，W，HICHW部CLOSELYRCLATCDT0MEPROLIFERATION鋤DDI任打E加『TIATIONOFTUMOR，CELLAPOPTOSIS，HNMUNEESCAPE，GEILESISOFBLOODVESSELS，ANDIILVASIONMDMETASTASISTHISEXPCRIMENT仃AILS斷EDTHEP1撇IDWHICHWAS誦THSTAT3GENETOBOVINEM鋤M哪EPITLLELIALCELLSN啪UGHTHE1IPOFACT鋤INE2000A11DSTUDIEDNLE111EBI010百CALCHARACT鰣STICSOFBOVINEM鋤MARREPITHELIALCELLS，PUTA劬D鋤EN謝ST印SFORPRODUCING仃ANSGELLICA11IMALSANDCELLIMMORTALIZATION1、BOVINEM鋤儺ARY印ITHELIALCELLSWEREIS01ATED鋤DP戚矗EDBYUSINGCOLLAGENASEDIGESTIONOFTHEM鋤MARR西AILDTISSUEFORMEPFIMA巧CELL伽LTURE，ANDSUBSCQU鋤TLYUSING仃YPSINSELECTEDDIGESTIONOFME饑1衄司CELLSFORODLP嘶FICATIONMO訕0109ICAL0BSERVATIONREVEALEDMATT11ECULTLLRCDCELLSPOSSESSEDTLLETYPICALCHARACTCROF印ITHELIALCELLS2、STAT3SI朗ALTRANSDUCERANDACTIVATOROFTRANS嘶PTION3GENEWASAMPLIFIED舶MPOTB7PLASMID訛CHCONTAINEDHUM鋤STAT3GCLLECDNA矗孵NEILT，TLLEILINS叭EDMOPEGFPCLVECTORTOCONS仃UCTREC0MBINANTPLASMID3、WBCULTILREDTHEBOVINEMAMMA巧EPITHELIALCELLSUSINGDMEMWITH15％FBS，也EN竹ANS斷EDTHESTAT3GEILETOCELLSOF廿LISTYPEMROU曲NLELIPOFACT鋤INE2000ANER48H，WEEX鋤INCDTLLEEXPRESSIONOFEGFPUSILLGNUORESCENTMICROSCOPE，24HOURSAREREXPO則RETOTHERECOMBINANTPL砌IDTLLE乜眥SFECTIONRALEOFTHEBEVCELLSW部L6％，SCREELLEDⅡLECLONEUSING600U咖LG418，也％MAKEDTHEDONEPFOPAGANDAUSING300UG／MLG4184、D舐VEDRNA舶MPOSITIVECLONECDLSALLDDESI弘EDTHEP枷CULARPRIMERTOGOT0THEIMPCRST印ATLAST，MEPOSITIVECELLSSHOWTHEAIMBAND2332BP，THENEGATIVECELLSHAVENOTHESAMEBAND5、FLOWCYTOME缸YWASUSEDT0ANDIYZEMECELLCYCLES、MULTIPLICATIONCAPACITYAND

簡(jiǎn)介：中國(guó)農(nóng)業(yè)大學(xué)博士學(xué)位論文摘要中文摘要哺乳動(dòng)物的乳腺是唯一在動(dòng)物出生后可以多次再生、退化的器官。不同發(fā)育時(shí)期的動(dòng)物乳腺中都存在一定比例的干細(xì)胞，正是這些干細(xì)胞維持和保證了乳腺發(fā)育的特性。利用類乳腺干細(xì)胞再生乳腺是近兩年來(lái)興起的一項(xiàng)高新技術(shù)，具有以一下四個(gè)方面的重要意義為那些因乳腺腫瘤或癌癥而切除乳腺的婦女修復(fù)或再生功能性乳腺為那些需要隆胸的女性提供安全可行的填充新材料為將健康成年雌性家畜乳腺改造成為分泌藥用或保健用蛋白的乳腺提供一個(gè)實(shí)驗(yàn)動(dòng)物的技術(shù)平臺(tái)為其他器官的發(fā)育、再生或修復(fù)提供方法、思路和理論借鑒我們以小鼠為模型，運(yùn)用組織化學(xué)、免疫熒光組織細(xì)胞化學(xué)、流式細(xì)胞儀分選方法FAGS以及分子生物學(xué)手段，研究了小鼠乳腺的發(fā)育規(guī)律小鼠乳腺織織中類乳腺干細(xì)胞小鼠乳腺細(xì)胞的分離、培養(yǎng)以及類乳腺干細(xì)胞的鑒定小鼠類乳腺干細(xì)胞分化的潛能小鼠乳腺類腺體體外短期培養(yǎng)富集類乳腺干細(xì)胞體系的優(yōu)化等。研究結(jié)果表明L3周齡的BALBC小鼠是去除乳腺腺體的最佳時(shí)期2不同發(fā)育時(shí)期乳腺組織中都存在類乳腺干細(xì)胞368周齡和懷孕期小鼠是獲得大量類乳腺干細(xì)胞的理想時(shí)期4首次證明體外短期培養(yǎng)的乳腺類腺體體系中，有一些細(xì)胞可以表達(dá)一定量的胚胎千細(xì)胞的特異性表達(dá)抗原基因OCT4REX1上皮干細(xì)胞的特異性表達(dá)抗原基因INTEGRIN31大部分已知干細(xì)胞共表達(dá)的抗原基因BCRPILACBG215小鼠類乳腺干細(xì)胞移植到去除乳腺腺體的小鼠乳脂墊中，可以再生功能性乳腺，因此，類乳腺干細(xì)胞在體內(nèi)具有分化潛能6小鼠類乳腺干細(xì)胞在體外誘導(dǎo)泌乳體系中培養(yǎng)7犬，可以表達(dá)P一酪蛋自及其基因，表明類乳腺干細(xì)胞在體外具有分化潛能7首次建立了優(yōu)化的乳腺類腺體體外短期培養(yǎng)富集類乳腺干細(xì)胞體系，由此體系培養(yǎng)的類腺體獲得類乳腺干細(xì)胞SEAL1的比例遠(yuǎn)遠(yuǎn)高于文獻(xiàn)報(bào)道使用體系獲得類乳腺干細(xì)胞比例8小鼠乳腺類腺體結(jié)構(gòu)體外短期培養(yǎng)獲得的細(xì)胞中有一定量的類乳腺干細(xì)胞存在優(yōu)化體系培養(yǎng)22小時(shí)獲得的細(xì)胞的類乳腺干細(xì)胞SEAL1比例遠(yuǎn)遠(yuǎn)高于培養(yǎng)72小時(shí)獲得細(xì)胞的比例相反，表達(dá)造血干細(xì)胞特異性抗原CD34的細(xì)胞CD34十比例在培養(yǎng)72小時(shí)比培養(yǎng)22小時(shí)高同時(shí)，培養(yǎng)的體系中還有一定比例的SEAL1CD34細(xì)胞存在。總之，隨著對(duì)類乳腺干細(xì)胞研究的深入，我們急需找到類乳腺干細(xì)胞的特異性表達(dá)抗原。在此之前，仍然只能以造血干細(xì)胞的特異性表達(dá)抗原SEALI作為一個(gè)標(biāo)記，進(jìn)行類乳腺干細(xì)胞的分析和分選研究。進(jìn)一步研究類乳腺干細(xì)胞的特異性表達(dá)譜，以期能夠找到類乳腺干細(xì)胞的特異性表達(dá)抗原，分離得到類乳腺干細(xì)胞，作為發(fā)育生物學(xué)和再生生物學(xué)研究的模FO另外，如果能對(duì)乳腺類腺體體外短期培養(yǎng)富集類乳IW干細(xì)胞體系進(jìn)行優(yōu)化，或許可以摸索適合類乳腺干細(xì)胞體外擴(kuò)增的培養(yǎng)體系。關(guān)鍵詞小鼠乳腺類乳腺干細(xì)胞SEALLCYTEKERATIN分化培養(yǎng)流式細(xì)胞儀中國(guó)農(nóng)業(yè)大學(xué)博士學(xué)位論文八T3STRACTINSUMMARYWESHOULDINVESTIGATETHEMOREEFFICIENTCULTURALSYSTEMFORMAMMARYORGNOIDSINVITROFORENRICHEDMOREMAMMARYEPITHELIALSTEMCELLSANDTOSCREENTHEEXPRESSIONPROFILESINMAMMARYEPITHELIALSTEMCELLSTODISCOVERTHESPECIFICMARKERFORMAMMARYEPITHELIALSTEMCELLSINORDERTOUTILIZETHEMODELOFMAMMARYEPITHELIALSTEMCELLSTOREGENERATETRANSGENICMAMMARYGLANDSINCOWORSHEEPFORHUMANHEALTHTOUNCOVERTHEMECHANISMOFORGANOGENESISANDDIFERENTIATIONKEYWORDSMOUSEMAMMARYGLANDSTEMCELLSSCALLCYTOKERATINDIFERENTIATIONCULTUREFLOWCYTOMETRYANALYSISSORTINGVI

簡(jiǎn)介：碩士學(xué)位論文SIRNASIRNA干擾干擾AIB3AIB3對(duì)乳腺癌細(xì)胞對(duì)乳腺癌細(xì)胞MDAMDAMBMB231231侵襲和轉(zhuǎn)移的影響侵襲和轉(zhuǎn)移的影響碩士研究生畢磊導(dǎo)師張灝教授專業(yè)生物化學(xué)與分子生物學(xué)研究方向腫瘤的發(fā)生與轉(zhuǎn)移入學(xué)時(shí)間2009年9月中國(guó)汕頭二零一二年五月學(xué)位論文原創(chuàng)性聲明本論文是我個(gè)人在導(dǎo)師指導(dǎo)下進(jìn)行的工作研究及取得的研究成果。論文中除了特別加以標(biāo)注和致謝的地方外，不包含其他人或其它機(jī)構(gòu)已經(jīng)發(fā)表或撰寫(xiě)過(guò)的研究成果。對(duì)本文的研究做出貢獻(xiàn)的個(gè)人和集體，均已在論文中以明確方式標(biāo)明。本人完全意識(shí)到本聲明的法律責(zé)任由本人承擔(dān)。作者簽名日期年月日學(xué)位論文使用授權(quán)聲明本人授權(quán)汕頭大學(xué)保存本學(xué)位論文的電子和紙質(zhì)文檔，允許論文被查閱和借閱；學(xué)?？蓪⒈緦W(xué)位論文的全部或部分內(nèi)容編入有關(guān)數(shù)據(jù)庫(kù)進(jìn)行檢索，可以采用影印、縮印或其它復(fù)制手段保存和匯編論文；學(xué)校可以向國(guó)家有關(guān)部門(mén)或機(jī)構(gòu)送交論文并授權(quán)其保存、借閱或上網(wǎng)公布本學(xué)位論文的全部或部分內(nèi)容。對(duì)于保密的論文，按照保密的有關(guān)規(guī)定和程序處理。作者簽名導(dǎo)師簽名日期年月日日期年月日

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