Department of Genetics, University of Cambridge, Downing Street, Cambridge CB2 3EH, UK.
Wellcome Trust Cancer Research UK Gurdon Institute, University of Cambridge, Tennis Court Road, Cambridge CB2 1QN, UK.
Development. 2018 Jul 11;145(13):dev164640. doi: 10.1242/dev.164640.
Cell type-specific transcriptome analysis is an essential tool for understanding biological processes in which diverse types of cells are involved. Although cell isolation methods such as fluorescence-activated cell sorting (FACS) in combination with transcriptome analysis have widely been used so far, their time-consuming and harsh procedures limit their applications. Here, we report a novel metabolic RNA sequencing method, SLAM-ITseq, which metabolically labels RNA with 4-thiouracil in a specific cell type followed by detection through an RNA-seq-based method that specifically distinguishes the thiolated uridine by base conversion. This method has successfully identified the cell type-specific transcriptome in three different tissues: endothelial cells in brain, epithelial cells in intestine and adipocytes in white adipose tissue. As this method does not require isolation of cells or RNA prior to the transcriptomic analysis, SLAM-ITseq provides an easy yet accurate snapshot of the transcriptional state .
细胞类型特异性转录组分析是理解涉及多种类型细胞的生物学过程的重要工具。尽管迄今为止已经广泛使用了诸如荧光激活细胞分选 (FACS) 与转录组分析相结合的细胞分离方法，但它们耗时且苛刻的程序限制了它们的应用。在这里，我们报告了一种新颖的代谢 RNA 测序方法，SLAM-ITseq，它使用 4-硫代尿嘧啶在特定细胞类型中代谢标记 RNA，然后通过一种基于 RNA-seq 的方法进行检测，该方法可以特异性地通过碱基转换区分硫代尿嘧啶。该方法已成功鉴定了三种不同组织中的细胞类型特异性转录组：脑内皮细胞、肠上皮细胞和白色脂肪组织中的脂肪细胞。由于这种方法在转录组分析之前不需要分离细胞或 RNA，因此 SLAM-ITseq 提供了细胞转录状态的简单而准确的快照。
