摘要
为构建瑟氏泰勒虫p23-IL-18复合基因真核表达质粒,以pMD-18T-p23和pMD-18T-IL-18克隆质粒为模板,应用重组PCR技术得到瑟氏泰勒虫p23-IL-18复合基因,将其先克隆到pMD-18-T载体,再亚克隆到真核表达载体pVAXⅠ中,得到重组质粒pVAXI-P23-IL-18。对该重组质粒进行PCR、酶切鉴定及测序后,采用脂质体法将其转染到BHK-21细胞,用RT-PCR和IFA进行鉴定。结果表明,瑟氏泰勒虫p23-IL-18复合基因真核表达质粒pVAXIP23-IL-18构建成功,可在BHK-21细胞中表达。本试验为p23-IL-18复合基因的免疫学研究奠定了基础。
Two pair of primers were designed by Oligo.6according to the sequence reported p23 gene of Theileria sergenti and IL-18 gene of cattle in the GenBank.Theileria sergenti p23 gene was linked with cattle IL-18 gene by SOE-PCR technique and then inserted into pMD18-T simple vector.The positive plasmid was digested with restriction-endonuclease of BamHI and EcoRI,and then the fusion gene was inserted into the eukaryotic expression vector pVAX1,named as pVAXp23-IL-18,the plasmid was transfected into BHK-21 cells with liposome,the expression of the fusion protein were analyzed by RT-PCR and IFA.The result indicated that pVAX-p23-IL-18 had been transfected into HeLa cells and transiently expressed successfully,which established the foundation for further study of the IL-18 and p23recombination gene as elect component of vaccinum.
出处
《中国兽医学报》
CAS
CSCD
北大核心
2015年第6期922-925,共4页
Chinese Journal of Veterinary Science
基金
延边大学"211工程"三期重点学科建设资助项目(2009-003)
