摘要
本实验以微小隐孢子虫 Cryptosporidium parvum 子孢子抗原P23基因和信号肽基因Usp45为材料,构建干酪乳杆菌 Lactobacillus casei 中分泌表达P23蛋白的重组载体pMG-Usp45-P23.在培养物上清和细菌裂解组分中,分别以SDS-PAGE、Western blotting、IFAT和ELISA等不同方法检测P23蛋白的分泌和表达情况,并用已获得的P23蛋白分泌产物体外刺激Wistar大鼠淋巴细胞,评价其淋巴细胞转化活性.结果表明,分别在细菌培养物上清和破碎菌体组分中检测到大小约为20 kDa（不含信号肽）和23 kDa （含信号肽）的P23蛋白分泌和表达产物.并且该P23蛋白分泌产物与ConA相当,具有显著的淋巴细胞转化活性.本实验为进一步研制动物微小隐孢子虫 C.parvum 活菌疫苗载体的构建奠定了基础.
In this study, the sporozoite antigen gene P23 and the signal peptide gene Usp45 of Cryptosporidium parvttm were used to construct the recombinant vector pMG-Usp45-p23 so as to secretory express the P23 protein in recombinant Lactobacillus. The secretion and expression of antigen P23 gene was detected by SDS-PAGE, Western blotting, IFAT and ELISA analysis in culture supernatants and bacterial disruption components respectively. The obtained P23 protein was used to stimulate the lymphocytes from Wistar rats in vitro in order to evaluate the activity of lymphocyte transformation.. Results revealed that products of P23 protein yield approximately 20 kDa （without signal peptide） and 23 kDa （with signal peptide） which were detected in culture supernatants and bacterial disruption components respectively. And the secretory products of IY23 protein had similar significant activity of lymphocyte transformation as ConA. The results have laied foundation for the further research on the construction of the live vaccine vectors of C. parvum in animals.
出处
《寄生虫与医学昆虫学报》
CAS
2012年第1期1-7,共7页
Acta Parasitologica et Medica Entomologica Sinica
基金
国家自然科学基金资助项目（N..30960279）
内蒙古农业大学“中国博士后流动站”项目（No.57545）
内蒙古农业大学“博士启动基金”项目（No.BJ054）2）
关键词
干酪乳杆菌
微小隐孢子虫
P23蛋白
分泌表达
Lactobacillus casei
Cryptosporidium parvum
P23 protein
Secretory expression
