摘要
目的 克隆弓形虫ZS2及RH株SAG3表面抗原基因片段 ,并进行序列分析。方法 设计合成引物 ,从弓形虫ZS2、RH及ZS1株基因组DNA中分别特异扩增出编码SAG3抗原的基因片段。扩增的目的片段经纯化后用EcoRⅠ和BamHⅠ双酶切后 ,克隆到原核表达质粒 pGEX - 4T - 2中 ,转化入大肠杆菌JM10 9,用PCR初筛 ,将PCR扩增阳性的重组子用EcoRⅠ和BamHⅠ双酶切鉴定 ,并进行序列的测定。结果 从弓形虫ZS2、RH和ZS1株DNA中扩增出 1176bp的SAG3基因 ,构建重组质粒 pGEX - 4T - 2 -SAG3(pGEX -SAG3) ,酶切产物的大小分别与预期相符。 结论 成功地对弓形虫ZS2、RH和ZS1株SAG3基因进行体外扩增及构建原核表达重组质粒 pGEX -SAG3,并经酶切及序列分析所验证 ,为弓形虫SAG3表面抗原的表达、体外诊断研究做好准备。
Aim To clone and sequence the SAG3 gene fragment of RH,ZS2 strains of Toxoplasma gondii Methods The SAG3 gene fragments of RH,ZS2 and ZS1 strains of Toxoplasma gondii were amplified by PCR After purified,and digested with EcoRⅠ+ BamHⅠ,the gene fragment was cloned as a plasmid pGEX 4T 2 The recombinant plasmid pGEX 4T 2 SAG3,was transformed into E coli JM109 Positive clones were screened and identified by PCR technique and digestion with restriction enzyme The sequence of inserted SAG3 gene fragment was also determined Results The SAG3 gene fragment was specifically amplified by PCR technique The recombinant plasmid pGEX SAG3 was constructed and digested by EcoRⅠ+ BamHⅠ,the size of gene fragment was 1176 bp and in accordance with the expected one Sequence determination analysis showed that the gene was gene fragment coding SAG3 antigen Conclusion The recombinant plasmid containing SAG3 gene from Toxoplasma gondii has been constructed successfully That can make it convenient for SAG3 antigen express and dignosis
出处
《中国人兽共患病杂志》
CSCD
北大核心
2003年第3期28-30,54,共4页
Chinese Journal of Zoonoses
基金
国家自然科学基金 (NO 3 0 0 70 682 )
"2 11工程"重点学科建设基金
广东省首批自然科学团队基金资助
