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

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.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • Animals
  • Argonaute Proteins / metabolism
  • Binding Sites
  • Cloning, Molecular / methods*
  • Mice
  • Nuclease Protection Assays
  • RNA Processing, Post-Transcriptional*
  • RNA, Antisense / metabolism*
  • RNA, Double-Stranded / genetics
  • RNA, Double-Stranded / metabolism*
  • RNA, Small Nucleolar / chemistry
  • RNA, Small Nucleolar / metabolism*
  • Reverse Transcriptase Polymerase Chain Reaction
  • Ribonucleases


  • Argonaute Proteins
  • RNA, Antisense
  • RNA, Double-Stranded
  • RNA, Small Nucleolar
  • Ribonucleases