摘要
组织特异性表达Cre重组酶的转基因小鼠是进行组织特异性条件敲除研究的关键。采用PCR扩增大鼠胰岛素基因705bp启动子指导Cre在胰岛细胞中特异表达;同时采用改构的Cre重组酶基因,在其5'端添加有真核核糖体结合序列和核定位序列使Cre重组酶能穿越核膜在细胞核能发挥功能;同时,为了保证原核基因Cre能在真核系统顺利表达,在其3'端添加含内含子的人生长激素基因。构建的表达载体在去除原核序列后用显微注射方法建立转基因小鼠,在出生的27只仔鼠中,PCR检测共获得7只Cre整合阳性的转基因小鼠,整合率26%。这种Cre转基因小鼠与基因组上携带LoxP位点的条件基因打靶小鼠交配,在胰腺组织中可以检测到Cre介导的重组,表明Cre在转基因小鼠胰腺中有表达。
The transgenic mice that express Cre in a tissue specific manner is a powerful tool in study of condition-al gene knockout.705bp insulin promoter was achieved to target the expression of Cre only in pancreatic.The Cre gene has been modified,include adding the nuclear localization signal and the sequence for initiation by eukaryotic ribosomes in Cre5' terminal.Cre gene was linked to human growth factor gene,which may improve the expression of Cre in transgenic mice.This construct was microinjected into mouse embryos.In the27offspring,7mice were i-dentified as being transgene by PCR method,and the efficiency was26%.The Cre transgenic mice were crossed with a conditional gene targeting mice to check the Cre mediated recombination.The results of PCR analysis sug-gested that the Cre recombinase was expressed in pancreas and could mediate the recombination between the LoxP sites in vivo.
出处
《生物技术通讯》
CAS
2002年第6期415-419,共5页
Letters in Biotechnology
