Systematic identification of sense-antisense transcripts in mammalian cells
Systematic identification of sense-antisense transcripts in mammalian cells
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DOI：
10.1038/nbt925
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发表时间：
2004-01-01
影响因子：
46.9
通讯作者：
Sioud, M
中科院分区：
文献类型：
--
作者：
Rosok, O;Sioud, M
在原核生物中，已经检测到许多内源性反义RNA，并发现它们具有多种生物学功能(1，2)。在真核生物中已经发现了反义RNA(3-7)；然而，缺乏实验方法来鉴定细胞中重叠的转录本是更系统地鉴定反义RNA的障碍。在这里，我们开发了一种实验策略，允许系统地识别与其他转录本具有长互补区域的内源mRNAs。该方法被应用于人正常乳腺上皮细胞和乳腺癌细胞。通过各种技术(如Northern杂交、RT-PCR)对正义和反义转录本的存在进行实验验证，支持该方法的特异性。当反义RNA被特定靶向时，其相应的mRNA水平显著改变，这一结果与已识别的反义RNA的调节作用一致。
In prokaryotes, a number of endogenous antisense RNAs have been detected and found to exert various biological functions(1,2). In eukaryotes antisense RNAs have been found(3-7); however, a lack of experimental methodologies that permit the identification of overlapping transcripts in cells presents a barrier to a more systematic identification of antisense RNA. Here we have developed an experimental strategy that allows systematic identification of endogenous mRNAs with long complementary regions to other transcripts. The method was applied to human normal mammary epithelial and breast cancer cells. Experimental validation of the presence of the sense and antisense transcripts by various techniques (e. g., northern blots, RT-PCR) supports the specificity of the method. When the antisense RNAs were specifically targeted, their corresponding mRNA levels significantly altered, a result consistent with a regulatory role for the identified antisense RNAs.