Esumi Shigeyuki, Wu Sheng-Xi, Yanagawa Yuchio, Obata Kunihiko, Sugimoto Yukihiko, Tamamaki Nobuaki
Department of Morphological Neural Science, Graduate School of Medical Sciences, Kumamoto University, 1-1-1 Honjo, Kumamoto 860-8556, Japan.
Neurosci Res. 2008 Apr;60(4):439-51. doi: 10.1016/j.neures.2007.12.011. Epub 2008 Jan 6.
The mammalian central nervous system is populated with various types of neurons and glia. To investigate the functions and development of individual cells requires gene-expression analysis at the single-cell level. Here, we developed a microarray-based method for the gene-expression profiling of single cells and tested it for GABAergic neuron progenitors. Single GABAergic neuron progenitors were collected from the neocortex of GAD67-GFP knock-in mice by dissociation followed by the aspiration of GFP-positive cells. Complementary DNA from the single cells was amplified by a method in which Super SMART PCR and T7 RNA polymerase amplification were combined at a optimized condition. The cRNA was subjected to microarray hybridization and analysis, which yielded reliable and reproducible results.
哺乳动物的中枢神经系统由各种类型的神经元和神经胶质细胞组成。要研究单个细胞的功能和发育，需要在单细胞水平上进行基因表达分析。在此，我们开发了一种基于微阵列的单细胞基因表达谱分析方法，并将其应用于γ-氨基丁酸能（GABAergic）神经元祖细胞的检测。通过解离从GAD67-GFP基因敲入小鼠的新皮质中收集单个GABAergic神经元祖细胞，然后吸出绿色荧光蛋白（GFP）阳性细胞。单细胞的互补DNA（cDNA）通过一种在优化条件下将超级智能PCR（Super SMART PCR）和T7 RNA聚合酶扩增相结合的方法进行扩增。将所得的互补RNA（cRNA）进行微阵列杂交和分析，得到了可靠且可重复的结果。
