摘要
克隆林麝(Moschus.berezovskii)白介素-2(interleukin-2,IL-2)到酵母表达载体pPIC9K,SalⅠ酶切后转化毕赤酵母GS115,G418抗性筛选高拷贝克隆,再由酵母菌落PCR鉴定;阳性克隆用甲醇诱导表达,SDS-PAGE分析,结果在诱导至第6d的酵母上清浓缩液中,检测到与预测林麝IL-2分子量相接近的,分子量约为18Ku的诱导表达带;TRIzol提取表达期间的酵母总RNA,并通过RT-PCR扩增IL-2mRNA,发现表达期的重组酵母细胞中存在IL-2mRNA,而对照没有检出,表明林麝IL-2存在低水平的表达。
Moschus berezovskii IL-2 gene was ligated into the yeast expression vector pPIC9K, then was transformed into picha pastoris GS115 strain by electroporation. Recombinants were identified by yeast colony PCR, then were induced by methanol and detected by SDS-PAGE analysis, The results of SDS-PAGE showed that after induced by methanol for 6 days, the concentrated supernant gave an induced band of forest musk deer IL-2 near 18 Ku, which was identified by RT-PCR amplification of IL-2 mRNA in expressing yeast.
出处
《中国预防兽医学报》
CAS
CSCD
北大核心
2007年第1期36-41,共6页
Chinese Journal of Preventive Veterinary Medicine
基金
国家科技部(项目编号:2003CCA0300)项目资助
关键词
林麝IL-2
毕赤酵母
真核表达
酵母菌落PCR
moschus berezovskii IL-2
picha pastoris
eukaryotic expression
yeast colony PCR
