华东理工大学学报：社会科学版
華東理工大學學報：社會科學版
화동리공대학학보：사회과학판
SOCIAL SCIENCES JOURNAL OF ECUST
2006年
8期
~
,共null页
口蹄疫病毒 噬菌体-scFv 可溶性scFv 噬菌体展示技术
口蹄疫病毒 噬菌體-scFv 可溶性scFv 噬菌體展示技術
구제역병독 서균체-scFv 가용성scFv 서균체전시기술
foot-and-mouth disease virus; phage-scFv; soluble scFv; phage displaying technology;
利用重组DNA技术在抗口蹄疫病毒单克隆抗体1C 7 VH基因和VL基因之间导入一段连接肽[(G ly4Ser)3],采用重叠延伸拼接法,经聚合酶链反应(PCR)扩增获得scF v基因。将scF v基因克隆至噬菌粒pCANTAB 5E载体,转化E.coli TG 1,构建噬菌体抗体文库。用M 13KO 7辅助噬菌体挽救及固相口蹄疫病毒(FMDV)抗原对噬菌体抗体文库的三轮“吸附-洗脱-扩增”的淘洗,筛选出scFv阳性克隆。将阳性克隆转化E.coli BH 2151,通过异丙基硫代-β-D-半乳糖苷(IPTG)诱导可溶性scFv蛋白的表达。酶联免疫吸附实验(EL ISA)检测表明:scFv克隆表达的phage-scFv及可溶性scFv与FMDV亲和力高,特异性强。
利用重組DNA技術在抗口蹄疫病毒單剋隆抗體1C 7 VH基因和VL基因之間導入一段連接肽[(G ly4Ser)3],採用重疊延伸拼接法,經聚閤酶鏈反應(PCR)擴增穫得scF v基因。將scF v基因剋隆至噬菌粒pCANTAB 5E載體,轉化E.coli TG 1,構建噬菌體抗體文庫。用M 13KO 7輔助噬菌體輓救及固相口蹄疫病毒(FMDV)抗原對噬菌體抗體文庫的三輪“吸附-洗脫-擴增”的淘洗,篩選齣scFv暘性剋隆。將暘性剋隆轉化E.coli BH 2151,通過異丙基硫代-β-D-半乳糖苷(IPTG)誘導可溶性scFv蛋白的錶達。酶聯免疫吸附實驗(EL ISA)檢測錶明:scFv剋隆錶達的phage-scFv及可溶性scFv與FMDV親和力高,特異性彊。
이용중조DNA기술재항구제역병독단극륭항체1C 7 VH기인화VL기인지간도입일단련접태[(G ly4Ser)3],채용중첩연신병접법,경취합매련반응(PCR)확증획득scF v기인。장scF v기인극륭지서균립pCANTAB 5E재체,전화E.coli TG 1,구건서균체항체문고。용M 13KO 7보조서균체만구급고상구제역병독(FMDV)항원대서균체항체문고적삼륜“흡부-세탈-확증”적도세,사선출scFv양성극륭。장양성극륭전화E.coli BH 2151,통과이병기류대-β-D-반유당감(IPTG)유도가용성scFv단백적표체。매련면역흡부실험(EL ISA)검측표명:scFv극륭표체적phage-scFv급가용성scFv여FMDV친화력고,특이성강。
The anti-foot-and-mouth disease virus(FMDV) single-chain Fv(scFv) gene was constructed by DNA recombinant technology.The 1C7V_H and V_L genes were assembled into scFv gene with a linker sequence(Gly_4Ser)_3 by splicing overlop extension and scFv gene was amplified by PCR.The scFv gene was subcloned into phagemid vector pCANTAB 5E.The phage scFv displaying library was constructed by male Escherichia coli TG1 transfected with resultant phagemid pCANTAB 5E-scFv.The scFv positive clone was obtained after the ph...
