Construction of Recombinant Adenovirus Expressing Protective Antigen of Bacillus Anthracis and Analysis of Immuno-responses

XU Lin; GUO Yan; GUAN Jie; YU Hong; ZHOU Yu-sen; LI Jin-song*

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June 2006

XU Lin, GUO Yan, GUAN Jie, YU Hong, ZHOU Yu-sen and LI Jin-song*. Construction of Recombinant Adenovirus Expressing Protective Antigen of Bacillus Anthracis and Analysis of Immuno-responses[J]. Virologica Sinica, 2006, 21(3): 231-234.

Citation: XU Lin, GUO Yan, GUAN Jie, YU Hong, ZHOU Yu-sen, LI Jin-song*. Construction of Recombinant Adenovirus Expressing Protective Antigen of Bacillus Anthracis and Analysis of Immuno-responses .VIROLOGICA SINICA, 2006, 21(3) : 231-234.

炭疽杆菌保护性抗原（PA）重组腺病毒的构建及免疫效果分析

1.
军事医学科学院微生物流行病研究所病原微生物生物安全国家重点实验室，北京，100071

摘要

探讨利用腺病毒载体作为炭疽杆菌基因工程疫苗载体的可行性。从载体pcDNA3.1-PA上PCR扩增PA片断，将该片断克隆入质粒pAdTrack-CMV，得到阳性克隆pAdTrack-PA。PmeI线性化的阳性克隆转化含有腺病毒骨架质粒pAdeasy-1的BJ5183感受态细胞，经同源重组后得到重组腺病毒vAd-PA。vAd-PA经PacI线性化后，脂质体介导转染293细胞，经Western- blot检测表明PA在293细胞中得到表达。重组病毒肌肉注射免疫BALB/c小鼠，用ELISA方法检测血清中产生了特异性抗体，抗体滴度计算几何均数为1：2800。该研究为进一步研究以腺病毒为活载体的疫苗奠定了基础。
- 炭疽杆菌
- , 保护性抗原
- , 重组腺病毒

Construction of Recombinant Adenovirus Expressing Protective Antigen of Bacillus Anthracis and Analysis of Immuno-responses

1.
State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing 100071，China

Abstract

In this study, we constructed recombinant adenovirus containing protective antigen gene of Bacillus anthracis. The PA gene was obtained from pcDNA3.1-PA vector by PCR , and cloned into plasmid pAdTrack-CMV. The positive clone was linearized with PmeI restriction enzyme and transformed into E. coli BJ5183 cells with the adenoviral plasmid pAdEasy-1. Then vAd-PA was generated by homologous recombination. After linearized with PacI restriction enzyme, vAd-PA was transfected into HEK-293 cell line by lipofection. Expression of PA in 293 cells was confirmed by SDS-PAGE and Western-blot. The BALB/c mice were intramuscularly inoculated with the recombinant adenovirus and the specific antibody was detected by ELISA . The induced antibody titer was 2.8×103 (Geometric Mean).
- Bacillus anthracis
- , Protective antigen
- , Recombinant adenovir

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通讯作者: 陈斌, bchen63@163.com

1.
沈阳化工大学材料科学与工程学院沈阳 110142

Construction of Recombinant Adenovirus Expressing Protective Antigen of Bacillus Anthracis and Analysis of Immuno-responses

1. State Key Laboratory of Pathogen and Biosecurity, Beijing Institute of Microbiology and Epidemiology, Beijing 100071，China

Abstract: In this study, we constructed recombinant adenovirus containing protective antigen gene of Bacillus anthracis. The PA gene was obtained from pcDNA3.1-PA vector by PCR , and cloned into plasmid pAdTrack-CMV. The positive clone was linearized with PmeI restriction enzyme and transformed into E. coli BJ5183 cells with the adenoviral plasmid pAdEasy-1. Then vAd-PA was generated by homologous recombination. After linearized with PacI restriction enzyme, vAd-PA was transfected into HEK-293 cell line by lipofection. Expression of PA in 293 cells was confirmed by SDS-PAGE and Western-blot. The BALB/c mice were intramuscularly inoculated with the recombinant adenovirus and the specific antibody was detected by ELISA . The induced antibody titer was 2.8×103 (Geometric Mean).