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| O型FMDV复合多表位在毕赤酵母中的表达与鉴定 | |
| 引用本文： | 胡 博,李 昌,王浩然,鲁会军,李 霄,朱战波,杜寿文,叶 明,袭 莹,韩佳丽,王 婧,金宁一.O型FMDV复合多表位在毕赤酵母中的表达与鉴定[J].中国兽医学报,2010,30(12). |
| 作者姓名： | 胡 博 李 昌 王浩然 鲁会军 李 霄 朱战波 杜寿文 叶 明 袭 莹 韩佳丽 王 婧 金宁一 |
| 作者单位： | 1. 吉林大学畜牧兽医学院，吉林长春130062；2．军事医学科学院军事兽医研究所全军基因工程重点实验室，吉林长春13006 2. 军事医学科学院军事兽医研究所全军基因工程重点实验室,吉林长春,130062 3. 吉林大学畜牧兽医学院,吉林长春,130062 4. 军事医学科学院军事兽医研究所全军基因工程重点实验室,吉林长春130062; 5. 吉林大学畜牧兽医学院，吉林长春130062；2．军事医学科学院军事兽医研究所全军基因工程重点实验室，吉林长春130062；3．黑龙江八一农垦大学动物科技学院，黑龙江大庆163319) 6. 吉林大学畜牧兽医学院，吉林长春130062；2．军事医学科学院军事兽医研究所全军基因工程重点实验室，吉林长春130062； 7. 军事医学科学院军事兽医研究所全军基因工程重点实验室,吉林长春130062;．黑龙江八一农垦大学动物科技学院，黑龙江大庆163319) |
| 摘 要： | 将O型FMDV复合多表位CTB-TEpi与FMDV结构基因P1-2A(P1/2A)连入毕赤酵母表达载体pPIC9K,经SacⅠ线性化后,利用电转化法整合到毕赤酵母(Pichia pastoris)基因组中。通过G418浓度梯度筛选得到高拷贝阳性转化子GS115/P1/2A-CTB-TEpi。经甲醇诱导表达后,目的蛋白在毕赤酵母中获得成功表达,并在FMDV自裂解片段2A的作用下裂解为P1/2A和CTB-TEpi 2个片段,相对分子质量分别为81 800和39 400,占上清液中可溶蛋白的比例为25%。Western blotting分析表明,表达蛋白的2部分均可被抗O型FMDV的血清所识别,而且含有CTB片段的CTB-TEpi条带可以被兔抗霍乱毒素血清识别,证明表达的蛋白具有抗原活性,为进一步研究O型FMDV复合多表位亚单位疫苗的免疫效果打下了基础。 |
| 关 键 词： | O型口蹄疫病毒 毕赤酵母 亚单位疫苗 |
Expression and identification of multi-epitope vaccine of FMDV serotype O in Pichia pastoris |
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| Hu Bo,LI Chang,W ANG Hao—ran,LU Hui—iun,LI Xiao,ZHU Zhan_ho,DU Shou—wen,YE M ing,xi ying,han jia li,wang jing,jia ning yi.Expression and identification of multi-epitope vaccine of FMDV serotype O in Pichia pastoris[J].Chinese Journal of Veterinary Science,2010,30(12). | |
| Authors: | Hu Bo LI Chang W ANG Hao—ran LU Hui—iun LI Xiao ZHU Zhan_ho DU Shou—wen YE M ing xi ying han jia li wang jing jia ning yi |
| Abstract: | Based on the Pichia pastoris secretory expression vector pPIC9K,a recombinant plasmids,pPIC9K-P1/2A-CTB-TEpi,containing capsid polypeptide P1-2A and multi-epitope of FMDV serotype O,have been constructed.Then the expression plasmid was transformed into P.pastoris GS115 by electroporation after lineared with Sac Ⅰ and integrated stably into chromosome of GS115.High-copied transformants GS115/P1/2A-CTB-TEpi was obtained by screen of G418 concentration gradient.Interest protein have been expressed successfully after induced by methanol,and clearaged two fragments,P1/2A and CTB-TEpi,as the autoclasia fragment 2A of FMDV.Molecular weight of the two fragments were 81 800 and 39 400,which accounted for about 25% of the total supernatant protein.The result of Western blotting showed that the two parts of the expression protein could be specifically recognited by serum against FMDV serotype O,and the fragment which contained CTB could be specifically recognited by serum against CT.This work provided a foundation for further study on the evaluation of the multi-epitope subunit vaccine of FMDV serotype O. |
| Keywords: | FMDV serotype O Pichia pastoris subunit vaccine |
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