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目的构建颞叶癫痫大鼠海马组织差异表达基因的消减cDNA文库,筛选颞叶癫痫的差异基因。方法采用抑制消减杂交方法,以正常大鼠和癫痫大鼠的海马组织作为对比材料,分离组织中差异表达基因的cDNA片段,将其与T载体进行T/A连接构建文库,将连接产物转化大肠杆菌进行文库扩增后,随机挑取100个白色克隆,用菌落PCR法筛选差异基因克隆,并进行测序分析。结果获得14个在癫痫大鼠海马组织中有差异表达,并与大鼠的已知基因片段高度同源的基因。结论抑制消减杂交技术是一种高效的筛选差异表达基因的方法,颞叶癫痫大鼠海马组织中存在许多差异表达基因,为深入研究癫痫发病的分子机制提供重要线索。
Objective To construct a subtractive cDNA library of differentially expressed genes in the temporal lobe epilepsy rat hippocampus to screen differentially expressed genes of temporal lobe epilepsy. Methods Suppression subtractive hybridization (SSH) method was used to isolate the cDNA fragments of differentially expressed genes from normal rats and hippocampus of epileptic rats. T-A cDNA was ligated with T vector to construct a library. The ligation products were transformed into large intestine After the library was amplified by the bacillus, 100 white clones were randomly picked, and the differential gene clones were screened by colony PCR and sequenced. Results Fourteen genes were found that were differentially expressed in the hippocampus of epileptic rats and highly homologous to the known gene fragments in rats. Conclusion Suppression subtractive hybridization is an efficient method for screening differentially expressed genes. There are many differentially expressed genes in the hippocampus of temporal lobe epilepsy rats, which provide important clues for further study on the molecular mechanism of epilepsy.