摘要
目的 从癫痫模型大鼠海马凋亡神经元中克隆caspase - 3的新底物。 方法 制作癫痫模型大鼠海马组织cDNA文库 ;PCR获得caspase- 3的P12和P17两亚基 ,然后分别定向插入pBridge质粒 ,构建三杂交诱饵载体 ;进行酵母三杂交筛库。结果 建立成功癫痫模型大鼠海马组织cDNA文库 ,构建了大鼠caspase- 3基因的酵母三杂交诱饵载体 ,并通过了caspase - 3与eIF2α之间的相互作用验证 ,筛库获得caspase- 3的新底物PIAS1。 结论 用酵母三杂交系统寻找caspase- 3下游底物具有可行性 ,从癫痫模型大鼠海马的cDNA文库中筛库获得caspase - 3的新底物PIAS1,为进一步研究caspase
Objective To clone the novel substrates of caspase-3 from the hippocampus apoptotic neurons of epileptic model rats. Methods Construct the yeast three-hybrid cDNA library with hippocampus of epileptic model rat. To get P12 and P17 subunits of caspase-3 through PCR from the library. Clone the two fragments into the Multiclone Sites (MCS) of pBridge vector for using yeast three-hybrid system to screen the caspase-3 interacting protein. Verify the feasibility of the bait vector with eIF2α. Results Constructing the yeast hybrid cDNA library with hippocampus of epileptic model rat successfully, construction of the bait vector for caspase-3 in yeast three-hybrid system was accomplished. Obtained a novel substrate of caspase-3,PIAS1. Conclusions The bait vector is feasible. With the bait we verify that caspase-3 can interact with eIF2α. Obtained PIAS1 as a novel substrate of caspase-3 in the hippocampus of epileptic model rat. PIAS1 may play certain role in neuronal apoptosis after status epilepsy.
出处
《神经疾病与精神卫生》
2004年第5期331-334,共4页
Journal of Neuroscience and Mental Health
