应用抑制性消减杂交技术筛选及鉴定高温致金黄地鼠神经管畸形下调表达基因

目的克隆高温致金黄地鼠神经管畸形下调表达基因，探讨其分子生物学机理。方法应用抑制性消减杂交技术构建高温致畸金黄地鼠胚胎神经管反向消减cDNA文库；联合蓝白斑和菌落PCR方法筛选阳性克隆进行测序和同源性分析；用Northern印迹方法证实这些基因的表达。结果成功构建高温致畸金黄地鼠胚胎神经管反向消减cDNA文库，从中筛选到15个基因，均与GenBank中已知基因有同源性。Northern印迹证实这些基因在高温致畸胚胎神经管中均呈下调表达。结论发现了高温致金黄地鼠神经管畸形过程中的一些重要相关基因并对其功能进行了初步探讨。

Cloning and characterization of down-regulated genes in neural tube defects of golden hamster induced by hyperthermia

OBJECTIVE: To clone down-regulated genes in neural tube defects of golden hamster induced by hyperthermia and to explore the molecular mechanism. METHODS: A reversed subtractive cDNA library was constructed using suppression subtractive hybridization. Clones with inserts were selected by combination of alpha complementary phenomenon and colony PCR. Then sequence and homology analysis of the inserts were made. And mRNA expression conditions were confirmed by Northern blot. RESULTS: The reversed subtractive cDNA library was successfully constructed. Fifteen recombinant clones were sequenced and recognized homologous to known genes. The results of Northern blot showed that all these genes were down-regulated in defected neural tubes induced by hyperthermia compared to normal developed neural tubes. CONCLUSION: Some important genes are identified which might be involved in neural tube defects induced by hyperthermia.