Comprehensive qPCR profiling of gene expression in single neuronal cells.
Comprehensive qPCR profiling of gene expression in single neuronal cells.
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DOI：
10.1038/nprot.2011.430
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发表时间：
2011-12-22
期刊：
影响因子：
14.8
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中科院分区：
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神经干细胞生物学中的一个主要挑战在于表征特定谱系的重新编程的人类神经细胞，这一过程需要使用对单细胞水平敏感的分析方法。单细胞基因图谱可以在高分辨率下提供关于一种细胞类型转化为另一种细胞类型的确切证据。我们描述的方法使用Fluidigm BioMark动态阵列从单个神经细胞高通量表达谱分析，在一个实验中分析多达96个独立样本和多达96个qPCR探针(相当于9216个反应)，该实验可以在2-3天内完成。该协议能够简单且经济高效地分析来自单个细胞的数百份转录本，并可能具有许多实用程序。
A major challenge in neuronal stem cell biology lies in characterization of lineage-specific reprogrammed human neuronal cells, a process that necessitates the use of an assay sensitive to the single-cell level. Single-cell gene profiling can provide definitive evidence regarding the conversion of one cell type into another at a high level of resolution. The protocol we describe employs Fluidigm Biomark dynamic arrays for high-throughput expression profiling from single neuronal cells, assaying up to 96 independent samples with up to 96 qPCR probes (equivalent to 9216 reactions) in a single experiment, which can be completed within 2–3 days. The protocol enables simple and cost-effective profiling of several hundred transcripts from a single cell, and could have numerous utilities.