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肿瘤相关抗原17-1A在毕赤酵母中的表达与鉴定
引用本文:王永生,田聆,刁鹏,胡敏,杜小波,魏于全. 肿瘤相关抗原17-1A在毕赤酵母中的表达与鉴定[J]. 四川大学学报(医学版), 2006, 37(4): 506-509
作者姓名:王永生  田聆  刁鹏  胡敏  杜小波  魏于全
作者单位:四川大学华西医院,生物治疗国家重点实验室,肿瘤中心,成都,610041;四川大学华西医院,生物治疗国家重点实验室,肿瘤中心,成都,610041;四川大学华西医院,生物治疗国家重点实验室,肿瘤中心,成都,610041;四川大学华西医院,生物治疗国家重点实验室,肿瘤中心,成都,610041;四川大学华西医院,生物治疗国家重点实验室,肿瘤中心,成都,610041;四川大学华西医院,生物治疗国家重点实验室,肿瘤中心,成都,610041
摘    要:目的利用Pichia pastoris酵母表达系统表达糖基化的肿瘤相关抗原17-1A,为进一步设计肿瘤蛋白疫苗提供研究基础。方法通过RT-PCR从小鼠肾组织中扩增17-1A的cDNA,将其定向克隆到pPICZαA质粒上,获得重组质粒pPICZαA-17-1A,测序正确后,重组质粒电转化到Pichia pastoris酵母菌株GS115中,在甲醇诱导下,利用其AOX I基因的α信号肽,分泌表达17-1A抗原糖蛋白,并对获得的蛋白进行SDS-PAGE及Western blot分析鉴定。结果构建了pPICZαA-17-1A重组质粒,通过电转化将目的基因整合人酵母基因组中,重组毕赤酵母表达能够表达17-1A抗原并检测到抗原发生了糖基化。结论能够通过毕赤酵母表达系统稳定表达糖基化的肿瘤相关抗原17-1A。

关 键 词:肿瘤相关抗原  17-1A  蛋白表达  毕赤巴斯德酵母  肿瘤疫苗
收稿时间:2005-09-06
修稿时间:2006-01-05

Expression and Characterization of Mouse 17-1A Antigen from Pichia pastoris
WANG Yong-sheng,TIAN Ling,DIAO Peng,HU Min,DU Xiao-bo,WEI Yu-quan. Expression and Characterization of Mouse 17-1A Antigen from Pichia pastoris[J]. Journal of Sichuan University. Medical science edition, 2006, 37(4): 506-509
Authors:WANG Yong-sheng  TIAN Ling  DIAO Peng  HU Min  DU Xiao-bo  WEI Yu-quan
Affiliation:State Key Laboratory of Biotherapy, Cancer Center, West China Hospital, Sichuan University, Chengdu 610041, China.
Abstract:OBJECTIVE: This is a ground work in preparation for the development of a vaccine against tumor, in which 17-1A glucoprotein, a tumor associated antigen (TAA), was cloned and expressed in Pichia pastoris. METHODS: The cDNA of 17-1A gene was cloned from nephridial tissue of mouse embryo by RT-PCR and subsequently inserted into pPICZaA plasmid after digestion with Xho I and Xba I. The linearized recombinant plasmid was transformed into Pichia pastoris strain GS115. 17-1A glucoproteins were secreted into culture supernatant through alpha-signal peptide of AOX I after inducation with methanol. The protein products were analyzed and characterized by SDS-PAGE and Western-blot. RESULTS: The plasmid pPICZalphaA-17-1A was constructed and verified by sequenceing. 17-1A glucoproteins were expressed stably and glycosylated in Pichia pastoris. CONCLUSION: Glycosylated tumor associated antigen 17-1A can be expressed in Pichia pastoris.
Keywords:17-1A
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