mRNA-Seq Whole Transcriptome Analysis of a Single Cell

Here we modified a single cell whole transcriptome amplification method to make it capable of amplifying cDNAs as long as 3kb efficiently and unbiasedly. We combined this modified single cell cDNA amplification method with Applied Biosystems next generation sequencing SOLiD System to set up a single cell whole transcriptome assay. The modified amplification strategy allows us to amplify full-length cDNAs for most of the expressed genes. We show that it is feasible to get digital gene expression profiles at single cell resolution. This allows us to ask fundamental biological questions that could not be addressed previously, especially in the early embryonic development field, and to understand transcriptome complexity at the resolution of a single cell. gene expression profiling from two single wild-type oocytes, two single Dicer knockout oocyte, and one single Ago2 knockout oocyte本研究对单细胞全转录组扩增方法进行改造，使其能够高效、无偏地扩增长达3kb的互补脱氧核糖核酸（cDNA）。本研究将改造后的单细胞cDNA扩增方法与应用生物系统公司（Applied Biosystems）下一代测序SOLiD系统相结合，构建了一套单细胞全转录组检测方法。该改造后的扩增策略可实现绝大多数表达基因的全长cDNA扩增。研究证实，在单细胞分辨率下获取数字基因表达谱具有可行性。这为我们解答此前难以攻克的核心生物学问题（尤其是早期胚胎发育领域）提供了可能，同时能够以单细胞分辨率解析转录组的复杂性。本数据集包含2个野生型卵母细胞、2个Dicer基因敲除卵母细胞及1个Ago2基因敲除卵母细胞的基因表达谱数据。