鸡堆型艾美耳球虫裂殖子单链抗体可变区基因文库的构建-预防兽医专业论文.docxVIP

The The Construction ofVariable Regions ofscFv LibraryAgainst the Merozoite of Eimeria acervullan AuthoEOuo“ AdvescE．Prof．Qin Jian-han Maj％Preventive Vetednmy Medicine Abstraet Four sli-ains Eimeria oocysts were obtained by the mothod ofsingle Oocyst isolation andⅥ*m studied outtobeEimeriaacercalian andEimeriatenella atfast．Atierpurification andextractiongenome DNA,by PCR amplification using species-spe啪c pfime堪,the four s仃ains Eimeria were：classified monuspecific Eimeria acervulian and Eimeria tenella． Eimeria acervulina merozoites w∞abstained horn intustinal muc嘲雠a methed sa咖out from four methods for farification of Eimerria口c们1f胁m merozmt髓．totaI RNA wa．q extracted击∞m the spleen cells ofBALB／C mice,which we犯immuned with merozoites soluble aetigon ofE．ac日、，ⅡliIla by four times，and immune咒spo咀s岱was measured by indirect ELISA． The frist chain of cDNA which derived from the total RNA with Obgo(dT)15Primer by RT WaS used tomplate,VH and VL genes w∞mnplified by PCR using gone specific primelⅢ．PCR preded be examined by agaruse gel electrophoresis and praised bY agarose gel DNA pufificafia kit，the result shows the genes of VH and VL amplified WCfe about 420bp and 390bp respectively．The l,mi6ed pruduct W88 cloned PGM-T vector．The cloned plasmids w∞transformed into TOP 10． Colonies were selected Oll ampicillin-onntaining agar plates and plasmids WCae isolated fr．om several independent clones．Thc clonies wcrl，sequenced by Sangers dideoxy sequencing method． Nucleotide sequence ofthe Vii and VL genes sequenced WCⅢe compared with the published VHand VL seqllell∞from the BALB／C mice．As expected,s既]uclice differe雠bdw嘲VH／VL and corresponding germline genes w∽identified．predominantly localized in complementary determining regions． Keywords：E．acervulina；merozoitcs；variable region of heavy chain唧)．variable region of light chain fvL)；∞Fv lilmⅡy 独创性声明本人声明所呈交的学位论文是本人在导师指导下进行的研究工作及取得的研究 独创性声明 本人声明所呈交的学位论文是本人在导师指导下进行的研究工作及取得的研究 成果。据我所知，除了文中特别加以标注和致谢的地方外，论文中不包含其他人已经 发表或撰写过的研究成果，也不包