CAJ | 학술논문

目的构建抗人结缔组织生长因子(CTGF)单链抗体(scFv)噬菌体展示文库.方法用重组人CTGF(rhCTGF)免疫balb/c小鼠,表达CTGF抗体后取小鼠脾脏获得B细胞,提取总RNA后反转录为cDNA,聚合酶链反应(PCR)获取重链可变区(vh)和轻链可变区(vl),vh和vl再次PCR装配为vl-linker-vh即scFv基因片段,并与噬菌粒PAK100酶连接,形成含有抗结缔组织生长因子单链抗体基因的目的噬菌粒；电转化入大肠杆菌XLl-blue,加入辅助噬菌体扩增后,建立噬菌体展示库重组库.结果经过凝胶电泳及噬菌粒基因测序分析,证实成功构建了抗结缔组织生长因子单链抗体定向免疫库.结论成功的构建了抗结缔组织生长因子单链抗体库,为制备抗结缔组织生长因子单链抗体奠定基础.
목적 구건항인결체조직생장인자(CTGF)단련항체(scFv)서균체전시문고.방법 용중조인CTGF(rhCTGF)면역balb/c소서,표체CTGF항체후취소서비장획득B세포,제취총RNA후반전록위cDNA,취합매련반응(PCR)획취중련가변구(vh)화경련가변구(vl),vh화vl재차PCR장배위vl-linker-vh즉scFv기인편단,병여서균립PAK100매련접,형성함유항결체조직생장인자단련항체기인적목적서균립；전전화입대장간균XLl-blue,가입보조서균체확증후,건립서균체전시고중조고.결과 경과응효전영급서균립기인측서분석,증실성공구건료항결체조직생장인자단련항체정향면역고.결론 성공적구건료항결체조직생장인자단련항체고,위제비항결체조직생장인자단련항체전정기출.
Objective To construct single chain Fv (scFv) library of human connective tissue growth factor (CTGF) by assembling the gene from immunized mice to phagemid PAK100.Methods Mice were immunized with recombinant human CTGF (rhCTGF).Total RNA was extracted from splenocytes of the immunized mice.Complementary DNA was got by reverse transcription PCR with the extracted RNA.Then the fragment of VH (variable heavy) and VL (variable light) were obtained by PCR.And they were assembled into scFv,which was inserted into phagemids vector PAK100 through SfiIsites.The phagemids containing scFV were transferred into E.coli.XL1-Blue bacterial cells to construct the scFv library to anti-human CTGF.Results Agarose gel clectrophoresis and sequences of phagemids certified that scFv phage display library to anti-human-CTGF was established.Conclusions The establishment of scFv library of anti-human CTGF set a foundation to get anti-human CTGF scFv.