ANALYSIS OF GENE-EXPRESSION IN SINGLE LIVE NEURONS
ANALYSIS OF GENE-EXPRESSION IN SINGLE LIVE NEURONS
复制标题
DOI：
10.1073/pnas.89.7.3010
复制
发表时间：
1992-04-01
影响因子：
11.1
通讯作者：
COLEMAN, P
中科院分区：
文献类型：
--
作者：
EBERWINE, J;YEH, H;COLEMAN, P
我们在这里提出了一种方法，广泛表征单细胞在分子水平上超越了更常见的形态和发射器/受体分类。 通过将引物、核苷酸和酶显微注射到来自大鼠脑的限定区域的急性解离的细胞中，扩增来自限定的单细胞的RNA。 进一步加工产生扩增的反义RNA。 第二轮扩增导致原始起始材料的> 10（6）倍扩增，这足以用于分析-例如，作为探针，cDNA文库的制备等。我们通过构建大鼠海马单个活细胞的表达谱来证明这种方法。 这种分析表明，似乎在形态上相似的细胞可能在表达模式上显示出明显的差异。 此外，我们从一个单一的细胞，其中一些是以前未描述的，可能是由于“稀有”时，平均在许多细胞类型的几个mRNA的特征。 电生理学分析与同一细胞内的分子生物学相结合，将有助于更好地理解分子水平上的变化如何表现为功能特性。 这种方法应该适用于各种各样的研究，包括发育，突变模型，衰老和神经退行性疾病。
We present here a method for broadly characterizing single cells at the molecular level beyond the more common morphological and transmitter/receptor classifications. The RNA from defined single cells is amplified by microinjecting primer, nucleotides, and enzyme into acutely dissociated cells from a defined region of rat brain. Further processing yields amplified antisense RNA. A second round of amplification results in > 10(6)-fold amplification of the original starting material, which is adequate for analysis-e.g., use as a probe, making of cDNA libraries, etc. We demonstrate this method by constructing expression profiles of single live cells from rat hippocampus. This profiling suggests that cells that appear to be morphologically similar may show marked differences in patterns of expression. In addition, we characterize several mRNAs from a single cell, some of which were previously undescribed, perhaps due to "rarity" when averaged over many cell types. Electrophysiological analysis coupled with molecular biology within the same cell will facilitate a better understanding of how changes at the molecular level are manifested in functional properties. This approach should be applicable to a wide variety of studies, including development, mutant models, aging, and neurodegenerative disease.