摘要
运用DNA重组技术将鸡白细胞介素2基因和鸡毒霉形体H3株TM-1基因进行串联,插入到pET-30a(+)质粒的EcoRⅠ和HindⅢ多克隆位点间,经PCR鉴定、双酶切鉴定和序列测定,表明已成功构建了含鸡毒霉形体H3株TM-1基因和鸡白细胞介素2基因的融合基因,将含有此融合基因的重组质粒命名为pET-30a(+)-TM-1-IL-2。将此重组质粒转化E.coliBL21(DE3)菌株。经IPTG 37℃诱导表达4 h后,SDS-PAGE电泳结果显示,此融合基因得到了表达,所表达出的融合蛋白的分子质量约为43 ku,主要以包涵体形式存在。表达产物在尿素存在下经超声波处理,获得了纯化的融合蛋白。这为进一步研究鸡白细胞介素2及鸡毒霉形体的生物学活性奠定了基础。
TM-1 gene from Mycoplasma gallisepticum （MG） H3 strain was linked with chicken IL-2 （ChIL-2） gene by DNA recombination technology and inserted into EcoR Ⅰ /Hind Ⅲ multiple clone site （MCS） of pET-30a（＋） plasmid as a fusion gene. PCR amplification, restriction digestion and sequencing confirmed that the recombinant plasmid containing the fusion gene was constructed, and designated pET- 30a（ ＋）-TM-1-IL-2. Then, pET-30a（ ＋ ）-TM-1-IL-2 was transformed into Escherichia coli BL21 （DE3）. SDS-PAGE analysis showed that the fusion gene was expressed efficiently in E. coli BL21（DE3） under induction of IPTG at 37 ℃ for 4 h and the expressed fusion protein was 43 ku in size as inclusion bodies. The fusion protein was purified by supersonic treatment with urea. The result provided a basis for further studies of the bioactivity of ChIL-2 and MG.
出处
《中国兽医科学》
CAS
CSCD
北大核心
2006年第12期967-971,共5页
Chinese Veterinary Science
基金
内蒙古农业大学博士科研启动基金项目(BJ04-4)
关键词
鸡毒霉形体
白细胞介素2
融合基因
原核表达
Mycoplasma gallisepticum
interleukin 2
fusion gene
prokaryotic expression
