Characterization of RNase R-digested cellular RNA source that consists of lariat and circular RNAs from pre-mRNA splicing

Nucleic Acids Res. 2006 May 8;34(8):e63. doi: 10.1093/nar/gkl151.


Besides linear RNAs, pre-mRNA splicing generates three forms of RNAs: lariat introns, Y-structure introns from trans-splicing, and circular exons through exon skipping. To study the persistence of excised introns in total cellular RNA, we used three Escherichia coli 3' to 5' exoribonucleases. Ribonuclease R (RNase R) thoroughly degrades the abundant linear RNAs and the Y-structure RNA, while preserving the loop portion of a lariat RNA. Ribonuclease II (RNase II) and polynucleotide phosphorylase (PNPase) also preserve the lariat loop, but are less efficient in degrading linear RNAs. RNase R digestion of the total RNA from human skeletal muscle generates an RNA pool consisting of lariat and circular RNAs. RT-PCR across the branch sites confirmed lariat RNAs and circular RNAs in the pool generated by constitutive and alternative splicing of the dystrophin pre-mRNA. Our results indicate that RNase R treatment can be used to construct an intronic cDNA library, in which majority of the intron lariats are represented. The highly specific activity of RNase R implies its ability to screen for rare intragenic trans-splicing in any target gene with a large background of cis-splicing. Further analysis of the intronic RNA pool from a specific tissue or cell will provide insights into the global profile of alternative splicing.

Publication types

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

MeSH terms

  • Dystrophin / genetics
  • Dystrophin / metabolism
  • Escherichia coli / enzymology
  • Escherichia coli Proteins / metabolism*
  • Exoribonucleases / metabolism*
  • Globins / genetics
  • Globins / metabolism
  • HeLa Cells
  • Humans
  • Introns
  • Nucleic Acid Conformation
  • Polyribonucleotide Nucleotidyltransferase / metabolism
  • RNA / chemistry
  • RNA / metabolism*
  • RNA Precursors / chemistry
  • RNA Precursors / metabolism*
  • RNA Splicing*
  • RNA, Circular
  • RNA, Messenger / chemistry
  • RNA, Messenger / metabolism*


  • Dystrophin
  • Escherichia coli Proteins
  • RNA Precursors
  • RNA, Circular
  • RNA, Messenger
  • rnr protein, E coli
  • RNA
  • Globins
  • Polyribonucleotide Nucleotidyltransferase
  • Exoribonucleases
  • exoribonuclease II