摘要
目的构建丝状支原体丝状亚种SC型LppQN端基因表达载体,并进行表达产物的纯化及免疫原性检测。方法利用PCR方法在体外定点突变丝状支原体丝状亚种SC型HVRIX株编码色氨酸的基因(TGA突变为TGG),构建表达载体,使其能在大肠杆菌中大量表达,并对表达产物进行纯化和免疫原性检测。结果表达产物约占菌体总蛋白的53.7%,经Westernblot和ELISA检测,表达蛋白具有良好的免疫原性。结论已成功表达了具有免疫原性的LppQ重组表达蛋白,可以作为诊断抗原用于血清学检测。
Objective To construct the expression vector of gene at N-terminus of lipoprotein LppQ of Mycoplasma mycoide subsp. Mycoides SC（ MmmSC）, purify the expressed product and study its immunogenicity. Methods The gene fragment encoding the N-terminal domain of LppQ was amplified from MmmSC HVRI-X strain by PCR then mutated from a Mycoplasma specific TGA（Trp） codon into a universal TGG （Trp） codon by one-step overlap extension PCR. The mutated gene was inserted into expression vector pE332a then transformed to E. coll. The expressed product was purified and determined for immunogenicity. Results The expressed product contained 53.7% of total somatic protein and showed good immunogenicity as proved by Western blot and ELISA. Conclusion Recombinant LppQ protein was successfully expressed and might be used as an antigen for serological diagnosis.
出处
《中国生物制品学杂志》
CAS
CSCD
2006年第3期278-281,共4页
Chinese Journal of Biologicals
基金
国家科技攻关计划项目(2002BA518A04).
关键词
丝状支原体丝状亚种SC型
脂蛋白
免疫原性
Mycoplasma mycoide subsp. Mycoides SC（MmmSC）
Lipoprotein
Immunogenicity
作者简介
通讯作者：辛九庆,Tel：0451-85935091,E-mail：xinjiuqing2001@sohu.com
