Search for genes involved in neuronal migration in cerebral cortex of X-irradiated embryonic mice

在前期低剂量电离辐射对啮齿类动物大脑皮层神经元迁移有抑制作用的基础上，我们尝试利用RLCS技术寻找与神经元迁移有关的基因RLCS的样品从胚胎14.5天的小鼠的全脑获得;一组经0.5戈伊X射线照射，另一组未经照射。迄今为止，我们用10种限制性内切酶对总共12，000个点进行了筛选。在比较来自辐射脑与来自非辐射脑的RLCS谱之后，我们发现在获得的点中，0.07%显示上调和0.11%下调。从显示下调的斑点中，我们成功地对其中一个下调的斑点进行了测序。该基因编码一种细胞粘附分子样蛋白，该蛋白由295至306个氨基酸组成，属于免疫球蛋白超家族，尽管该分子的功能尚不清楚。应用实时荧光定量PCR方法，证实了X射线照射后该基因的表达水平被抑制到未照射对照的30-40%，并尝试设计了一种新的方法，即“单细胞RLCS”，使我们能够在单细胞水平上研究基因表达的变化。在本研究计画期间，我们已完成单细胞RLCS的技术建立。

On the basis of our previous studies that prenatal low-level ionizing radiation impedes neuronal migration in rodent cerebral cortices, we have attempted to search for genes involved in neuronal migration by using RLCS (Restriction Landmark cDNA Scanning) method.The samples for RLCS were obtained from the whole brains of embryonic day 14.5 mice ; one group X-irradiated at 0.5 Gy, the other non-irradiated, We have so far made screening on total 12,000 spots by employing 10 restriction enzymes. After comparing the RLCS profiles from irradiated brains with those from non-irradiated brains we found that among the spots obtained 0.07% showed up-regulation and 0.11% down-regulation. From the spots showing down-regulation we succeeded in sequencing one of the down-regulated spots. The gene codes for one of the cell adhesion molecule like protein comprising of 295 to 306 amino acids, which belongs to the immunoglobulin superfamily, although the function of the molecule remains unknown. By applying real-time PCR method, we have confirmed that the expression level of the gene in X-irradiated brains was suppressed down to 30-40% of the non-irradiated control.We have also attempted to devise a new method, called "single cell RLCS" that enables us to study the changes of gene expression at single cell level. During the term of the present research project we have completed in establishing single cell RLCS from the technical viewpoint.