Alternative transcription start sites (TSSs) usage plays a critical role in gene transcription regulation in mammals. However, precisely identifying alternative TSSs remains challenging at the genome‐wide level. We report a single‐cell genomic technology for alternative TSSs annotation and cell heterogeneity detection. In the method, we utilize Fluidigm C1 system to capture individual cells of interest, SMARTer cDNA synthesis kit to recover full‐length cDNAs, then dual priming oligonucleotide system to specifically enrich TSSs for genomic analysis. We apply this method to a genome‐wide study of alternative TSSs identification in two different IFN‐β stimulated mouse embryonic fibroblasts (MEFs). The data clearly discriminate two IFN‐β stimulated MEFs. Moreover, our results indicate 81% expressed genes in these two cell types containing multiple TSSs, which is much higher than previous predictions based on Cap‐Analysis Gene Expression (CAGE) (58%) or empirical determination (54%) in various cell types. This indicates that alternative TSSs are more pervasive than expected and implies our strategy could position them at an unprecedented sensitivity. It would be helpful for elucidating their biological insights in future.

中文翻译：

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用于识别替代转录起始位点的单细胞基因组策略

替代转录起始位点（TSS）的使用在哺乳动物的基因转录调控中起着至关重要的作用。然而，在全基因组水平上精确识别替代 TSS 仍然具有挑战性。我们报告了一种用于替代 TSS 注释和细胞异质性检测的单细胞基因组技术。在该方法中，我们利用 Fluidigm C1 系统捕获感兴趣的单个细胞，利用 SMARTer cDNA 合成试剂盒回收全长 cDNA，然后利用双引发寡核苷酸系统特异性富集 TSS 以进行基因组分析。我们将此方法应用于两种不同 IFN-β 刺激的小鼠胚胎成纤维细胞 (MEF) 中替代 TSS 鉴定的全基因组研究。数据清楚地区分了两种 IFN-β 刺激的 MEF。此外，我们的结果表明，在这两种含有多个 TSS 的细胞类型中，有 81% 的基因表达，这远远高于之前基于 Cap 分析基因表达 (CAGE) (58%) 或各种细胞类型的经验确定 (54%) 的预测。这表明替代 TSS 比预期更普遍，并且意味着我们的策略可以将它们置于前所未有的敏感度。这将有助于将来阐明他们的生物学见解。