弓形虫速殖子噬菌体展示文库的构建  

Display Library Construction of Toxoplasma Tachyzoite Phage

作　　者：王春苗 卢致民 王燕 WANG Chun-miao;LU Zhi-min;WANG yan(Department of Pathogenic Biology,School of Medical Laboratory,Hebei North University,Zhangjiakou,Hebei 075000,China)

机构地区：[1]河北北方学院医学检验学院病原生物学教研室

出　　处：《河北北方学院学报（自然科学版）》2019年第12期1-4,共4页Journal of Hebei North University:Natural Science Edition

基　　金：河北省高等学校科学技术研究项目(No.Z2017025);河北北方学院创新人才培育项目(No.CXRC1326)

摘　　要：目的构建弓形虫速殖子λ噬菌体展示文库,为后期免疫筛选做准备。方法通过Trizol试剂提取弓形虫RH株速殖子总RNA,再利用SMART技术进行反转录合成双链cDNA。对cDNA进行纯化后,通过Sfi I酶切消化双链cDNA并切胶回收,与λTripIEx2载体的两臂连接,最后通过λ噬菌体包装蛋白进行体外包装,以构建λ噬菌体展示文库。对文库进行滴度测定,并进行重组率检测。结果RNA提取结果良好,未扩增文库滴度为8.0×10^6 pfu·mL^-1,文库重组率为95%。随机挑取50个噬菌斑进行PCR鉴定,插入片段分布在0.5~3 kb。结论成功构建了弓形虫速殖子λ噬菌体展示文库,为后期疫苗的研究奠定基础。Objective To construct aλphage display library of Toxoplasma tachyzoite,and to prepare for later immune-screening.Methods Total RNA was extracted from Toxoplasma RH strain tachyzoite by Trizol reagent,then the double-stranded cDNA was synthesized by reverse transcription through SMART technology.After purification of cDNA,the double strand cDNA was digested by SFI I enzyme and recovered by gel cutting,then connected to the two arms ofλTripIEx2 carrier.Finally,the packaging protein ofλphage was packed in vitro to constructλphage display library.The titer of cDNA library and recombination rate were measured.Results RNA extraction result was good.The titer of the unamplied library was 8.0×10^6 pfu·mL^-1,and the recombination rate of the library was 95%.50 phage plaques were randomly selected for PCR identification,and the inserted fragments were distributed in 0.5~3 kb.Conclusionsλphage display library of Toxoplasma tachyzoite is constructed successfully,which lays the foundation for research of later vaccines.

关 键 词：弓形虫速殖子 疫苗 λ噬菌体展示文库 

分 类 号：S85[农业科学—兽医学]