Roma Guglielmo, Cobellis Gilda, Claudiani Pamela, Maione Francesco, Cruz Pedro, Tripoli Gaetano, Sardiello Marco, Peluso Ivana, Stupka Elia
Telethon Institute of Genetics and Medicine, 80131 Napoli, Italy.
Genome Res. 2007 Jul;17(7):1051-60. doi: 10.1101/gr.5720807. Epub 2007 May 31.
Embryonic stem (ES) cells are pluripotent cell lines with the capacity of self-renewal and the ability to differentiate into specific cell types. We performed the first genome-wide analysis of the mouse ES cell transcriptome using approximately 250,000 gene trap sequence tags deposited in public databases. We unveiled >8000 novel transcripts, mostly non-coding, and >1000 novel alternative and often tissue-specific exons of known genes. Experimental verification of the expression of these genes and exons by RT-PCR yielded a 70% validation rate. A novel non-coding transcript within the set studied showed a highly specific pattern of expression by in situ hybridization. Our analysis also shows that the genome presents gene trapping hotspots, which correspond to 383 known and 87 novel genes. These "hypertrapped" genes show minimal overlap with previously published expression profiles of ES cells; however, we prove by real-time PCR that they are highly expressed in this cell type, thus potentially contributing to the phenotype of ES cells. Although gene trapping was initially devised as an insertional mutagenesis technique, our study demonstrates its impact on the discovery of a substantial and unprecedented portion of the transcriptome.
胚胎干细胞（ES细胞）是具有自我更新能力且能够分化为特定细胞类型的多能细胞系。我们利用公共数据库中存放的约250,000个基因捕获序列标签，首次对小鼠ES细胞转录组进行了全基因组分析。我们发现了8000多个新转录本，其中大部分是非编码的，还发现了1000多个已知基因的新的可变外显子，且这些外显子通常具有组织特异性。通过逆转录聚合酶链反应（RT-PCR）对这些基因和外显子的表达进行实验验证，验证率达70%。在所研究的一组新的非编码转录本中，有一个通过原位杂交显示出高度特异性的表达模式。我们的分析还表明，基因组中存在基因捕获热点，这些热点对应于383个已知基因和87个新基因。这些“高度捕获”的基因与先前发表的ES细胞表达谱几乎没有重叠；然而，我们通过实时PCR证明它们在这种细胞类型中高度表达，因此可能对ES细胞的表型有贡献。尽管基因捕获最初被设计为一种插入诱变技术，但我们的研究证明了它对发现转录组中大量前所未有的部分所产生的影响。
