title: Direct cloning of double-stranded RNAs from RNase protection analysis reveals processing patterns of C/D box snoRNAs and provides evidence for widespread antisense transcript expression.
abstract: 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.
authors: Department of Molecular and Cellular Biochemistry, University of Kentucky, Lexington, KY, 40536, USA.
authors: Manli
authors: M
authors: Shen
authors: Eduardo
authors: E
authors: Eyras
authors: Jie
authors: J
authors: Wu
authors: Amit
authors: A
authors: Khanna
authors: Serene
authors: S
authors: Josiah
authors: Mathieu
authors: M
authors: Rederstorff
authors: Michael Q
authors: MQ
authors: Zhang
authors: Stefan
authors: S
authors: Stamm
mesh_terms: D000818:Animals; D060565:Argonaute Proteins / Q000378:metabolism; D001665:Binding Sites; D003001:Cloning, Molecular / Q000379:methods*; D051379:Mice; D023282:Nuclease Protection Assays; D012323:RNA Processing, Post-Transcriptional; D016372:RNA, Antisense / Q000378:metabolism*; D012330:RNA, Double-Stranded / Q000235:genetics / Q000378:metabolism*; D020537:RNA, Small Nucleolar / Q000737:chemistry / Q000378:metabolism*; D020133:Reverse Transcriptase Polymerase Chain Reaction; D012260:Ribonucleases
chemical_list: D060565:Argonaute Proteins; D016372:RNA, Antisense; D012330:RNA, Double-Stranded; D020537:RNA, Small Nucleolar; D012260:Ribonucleases