Department of Molecular and Cellular Biochemistry, University of Kentucky, Lexington, KY, 40536, USA.
Nucleic Acids Res. 2011 Dec;39(22):9720-30. doi: 10.1093/nar/gkr684. Epub 2011 Aug 31.
We describe a new method that allows cloning of double-stranded RNAs (dsRNAs) that are generated in RNase protection experiments. We demonstrate that the mouse C/D box snoRNA MBII-85 (SNORD116) is processed into at least five shorter RNAs using processing sites near known functional elements of C/D box snoRNAs. Surprisingly, the majority of cloned RNAs from RNase protection experiments were derived from endogenous cellular RNA, indicating widespread antisense expression. The cloned dsRNAs could be mapped to genome areas that show RNA expression on both DNA strands and partially overlapped with experimentally determined argonaute-binding sites. The data suggest a conserved processing pattern for some C/D box snoRNAs and abundant expression of longer, non-coding RNAs in the cell that can potentially form dsRNAs.
我们描述了一种新的方法，该方法允许克隆在核糖核酸酶保护实验中产生的双链核糖核酸 (dsRNAs)。我们证明，小鼠 C/D 框 snoRNA MBII-85 (SNORD116) 使用 C/D 框 snoRNAs 的已知功能元件附近的加工位点被加工成至少五种较短的 RNA。令人惊讶的是，来自核糖核酸酶保护实验的大多数克隆 RNA 来源于内源性细胞 RNA，表明广泛的反义表达。克隆的 dsRNAs 可以映射到显示两条 DNA 链上 RNA 表达的基因组区域，并与实验确定的 Argonaute 结合位点部分重叠。这些数据表明，一些 C/D 框 snoRNAs 具有保守的加工模式，并且细胞中大量表达潜在能够形成 dsRNAs 的较长非编码 RNA。
