点击查看其他>>兽医硕士论文(优秀范文6篇)兽医硕士论文范文第三篇：本篇文章目录导航：[][第一章]牛繁殖关键技术探究[第二章]体外培养胚胎分泌外泌体的分离与鉴定[第三章]培养体外胚胎分泌外泌体对核移植胚胎发育的 更多精彩就在： 51免费论文网|www.jxszl.com 
影响[第四章-参考文献]不同量的外泌体对核移植胚胎囊胚率的影响结论摘要胚胎在体外培养的发育过程中,在形态.基因表达和发育能力方面不能够达到胚胎在雌性生殖道内的发育的水平,因此优化培养液成分仍然是非常迫切需要.牛胚胎体外培养时,需要培养7天发育到囊胚,然后进行胚胎移植.通常采用第3.5天更换胚胎培养液,然而更换培养液对胚胎的影响仍然不清楚.采用超速离心法提取胚胎培养液中的外泌体,采用投射电子显微镜和免疫荧光染色外泌体特异性蛋白CD9鉴定外泌体,发现胚胎在体外培养过程中能够分泌外泌体,换液会导致胚胎分泌外泌体丢失.用30μL新鲜培养液重悬从10mL第3.5天胚胎培养液提取的外泌体.将不同量(1μL.5μL和10μL)的胚胎外泌体提取物添加到胚胎培养液中,添加5μL和10μL外泌体提取物能够显着提高胚胎体外发育的囊胚率.将胚胎分为三组培养,分别是换液组(第3.5天换液).不换液组(全程不换液)和外泌体添加组(第3.5天换液并添加5μL外泌体提取物).基因表达结果表明换液影响胚胎发生内质网应激.凋亡和内细胞团和滋养层细胞分化.添加外泌组的Oct4基因的表达水平显着高于换液组,这表明换液导致丢失的胚胎分泌的外泌体在内细胞团和滋养层细胞分化过程中发挥重要的作用.本探究发现核移植胚胎体外培养过程中,换液导致胚胎外环境发生剧烈变化诱发胚胎发生内质网应激与凋亡,不利于胚胎发育；换液过程中导致胚胎分泌的外泌体丢失,影响核移植胚胎内细胞团和滋养层细胞分化；胚胎分泌的外泌体在调控胚胎体外发育中发挥重要的作用.本探究对优化胚胎的体外培养,提高克隆效率,为提高牛繁殖育种效率提供技术支撑.关键字：外泌体；胚胎；体细胞核移植；体外培养；发育ABSTRACTThedevelopmentalcompetenceofinvitrocultured(IVC)embryosismarkedlylowerthanthatoftheirinvivocounterparts,suggestingtheneedforoptimizationofIVCprotocols.Embryoculturemediumisroutinelyreplacedthreedaysafterinitialcultureinbovine,however,whetherthisprotocolissuperiortocontinuousnonrenewalculturemethodundercurrentconditionsremainsunclear.Exosomesinembryoculturemediumwereextractedbyultracentrifugation.TheexosomeswereidentifiedbyelectronmicroscopyandimmunofluorescencestainingwithexosomespecificproteinCD9.Theresultsshowedthattheexosomescouldbesecretedbytheembryoduringinvitroculture,andtheexosomeswouldbelostbychangingembryoculturemedium.Theexosomesextractedfrom10mLembryoculturemediumonday3.5weresuspendedin30uLfreshculturemedium.Adding5uLand10uLgroupsshowedsignificantlyhigherblastocystratesthanthecontrolgroupand1uL.Alsoembryosweredividedintothreegroups,renewalgroup(renewedon3.5days),non-renewalgroup(withoutrenewalduringIVC)andsupplementarygroup(supplementedexosomeson3.5daysduringrenewal).Theresultsofgeneexpressionshowedthatrenewalaffectedendoplasmicreticulumstress,apoptosisanddifferentiationofinnercellmassandtrophoblastcells.TheexpressionlevelofOct4geneinthesupplementarygroupwassignificantlyhigherthanthatinrenewalgroup,suggestingthatexosomessecretedbyembryosplayanimportantroleinthedevelopmentofinnercellmassandtrophoblastcells.Inthisstudy,wefoundthatrenewalinducedendoplasmicreticulumstressandapoptosis,whichwasnotconducivetoembryonicdevelopment;duringtheprocessofrenewal,theexosomessecretedbyembryoswerelost,whichaffectedthedifferentiationofinnercellmassandtrophoblastcells.Thisstudywillprovidetechnicalsupportforoptimizingembryocultureinvitro,improvingcloningefficiencyandimprovingreproductiveandbreedingefficiencyofcattle.KEYWORLD:Exosome;Embryo;Somaticcellnucleartransfer;invitrocultured;development目录第一章综述1.1牛繁殖关键技术探究进展1.1.1人工授精1.1.2超数排卵与胚胎收集1.1.3胚胎移植1.1.4胚胎培养1.1.5基因编辑技术1.1.6体细胞核移植1.2外泌体探究进展1.2.1外泌体的生理作用1.2.2外泌体在疾病发生中的作用1.2.3外泌体在诊断中的作用1.2.4外泌体在治疗中的作用1.2.5外泌体的分离纯化和储存第二章体外培养胚胎分泌外泌体的分离与鉴定2.1试验材料与方法2.1.1试验材料2.1.2试验方法2.2试验结果2.2.1核移植胚胎体外培养分泌外泌体的电镜结构2.2.2核移植胚胎体外培养分泌外泌体的免疫荧光检测2.3探讨2.4小结第三章体外培养胚胎分泌外泌体对核移植胚胎发育的影响3.1试验材料与方法3.1.1试验材料3.1.2试验方法3.2试验结果3.2.1不同量的外泌体对核移植胚胎囊胚率的影响3.2.2换液.不换液和添加外泌体对核移胚胎基因表达的影响3.2.3换液.不换液和添加外泌体对核移胚胎体外培养囊胚率的影响3.3探讨3.4小结结论参考文献
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