同时表达多种外源基因的非复制型重组痘苗病毒的构建  

CONSTRUCTION OF NON REPLICATING VACCINIA RECOMBINANT VIRUS EXPRESSING MULTIPLE FOREIGN GENES

作　　者：马海伦[1] 孙朝晖[1] 陆柔剑[1] 娄元梅[1] 郭斐[1] 张帆[1] 阮力[1] 

机构地区：[1]中国预防医学科学院病毒学研究所

出　　处：《病毒学报》1999年第1期21-28,共8页Chinese Journal of Virology

基　　金：国家863高科技生物领域及国家自然科学基金

摘　　要：利用非复制型痘苗病毒载体，构建了能同时表达乙型肝炎（乙肝）病毒ＳＳ１、麻疹病毒ＨＡ和Ｆ及白细胞介素２（ＩＬ２）的非复制型重组痘苗病毒ＶＨＦＩＬ２ΔＣＫＳＳ１，及其相应的复制型重组痘苗病毒ＶＨＦＩＬ２ＳＳ１。这两株重组病毒在ＣＥＦ细胞上连续传至第２５代，经Ｓｏｕｔｈｅｒｎｂｌｏｔ证实，两株病毒均在Ａ２４、Ａ２７间稳定整合有ＳＳ１基因，非复制型重组病毒Ｃ、Ｋ间基因稳定缺失。经ＲＩＡ、ＥＬＩＳＡ及Ｗｅｓｔｅｒｎｂｌｏｔ方法检测，这两株病毒均可稳定分泌表达ＳＳ１蛋白，其分泌表达的ＳＳ１蛋白与ＣＨＯ细胞表达的ＳＳ１蛋白性质相近，接近于天然。经免疫打点检测，两株病毒均可稳定表达麻疹病毒的ＨＡ与Ｆ蛋白以及ＩＬ２。而且ＶＨＦＩＬ２ΔＣＫＳＳ１与ＶＨＦＩＬ２ＳＳ１表达多种外源抗原的水平无显著性差异。ＶＨＦＩＬ２ΔＣＫＳＳ１经皮内注射免疫家兔，４周后可诱发抗乙肝病毒ＳＳ１、抗麻疹病毒及抗痘苗病毒的特异性ＩｇＧ抗体；加强免疫２周后，特异性抗体滴度可上升４～１６倍，与其初始病毒ＶＨＦＩＬ２ΔＣＫ诱发的抗体水平相同，比ＶＨＦＩＬ２ＳＳ１诱发的抗体滴度低０～４倍。非复制型多价重组痘苗病毒的构建，为评价非复制型痘苗病毒载体作为多价疫苗载?The non replicating recombinant VHFIL2ΔCKSS1 and replicating recombinant VHFIL2SS1 expressing SS1 protein of HBV, HA and F proteins of measles virus and IL 2 were constructed using non replicating vaccinia virus vector.By Southern blot,it was determined that the SS1 gene was integrated stably in both of the two recombinants after twenty five passages in CEF cell,and the genes between C and K fragments of the non replicating were deleted stably.Further investigation by RIA,ELISA and Western blot demonstrated that the two recombinants could stably express SS1 protein with natural properties,and was similar to that expressed in CHO cell.The dot ELISA results showed that the two recombinants could express stably HA and F proteins of measles virus and IL2. And the expression level of multiple foreign antigens expressed by VHFIL2ΔCKSS1 and VHFIL2SS1 did not make significant differences.By intradermal immunization,specific IgG antibody against SS1,measles and vaccinia viruses could be induced by VHFIL2ΔCKSS1 in rabbits after 4 weeks,and the IgG titer could be boosted 4～16 times after revaccination two weeks later,which reached a similar level to that induced by VHFIL2ΔCK but was 0 4 times lower than that induced by VHFIL2SS1.Construction of non replicating multivalent recombinant provides a base to evaluate efficacy and safety of non replicating vaccinia virus as a vector of multivalent vaccine.

关 键 词：多价疫苗 痘苗病毒载体 遗传稳定性 免疫效果 

分 类 号：R392[医药卫生—免疫学]