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. 1988 Jul 25;16(14A):6597–6605. doi: 10.1093/nar/16.14.6597

Discrimination between RNA circles, interlocked RNA circles and lariats using two-dimensional polyacrylamide gel electrophoresis.

H F Tabak 1, G Van der Horst 1, J Smit 1, A J Winter 1, Y Mul 1, M J Groot Koerkamp 1
PMCID: PMC338316  PMID: 2456529

Abstract

Two-dimensional polyacrylamide gel electrophoresis can be used to identify structural forms of RNA such as linear RNA, circular RNA, interlocked circles and lariats. The procedure is based upon the characteristic migration behaviour of the degradation products derived from the intact structures present already before the start of the experiment or formed during or after electrophoresis in the first dimension. After autoradiography to detect the positions of the radiolabeled RNA molecules, circles broken during electrophoresis of the first dimension give rise to horizontal lines touching the diagonal formed by linear RNAs at a point corresponding to the length of the RNA circle from which it was derived. Products derived from interlocked RNA circles by breakage after completion of the first dimension appear on a vertical line underneath the intact complex and consist of free RNA circles and their linear derivatives. Broken lariats give rise to two lines depending on the location of the break. Lariats with broken tails are present on a line to a position that corresponds to the length of their tail and that runs parallel to the diagonal formed by linear products. Lariats with a broken eye form a line running from the position of the intact product to the diagonal formed by the linear RNAs.

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Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Aaij C., Borst P. The gel electrophoresis of DNA. Biochim Biophys Acta. 1972 May 10;269(2):192–200. doi: 10.1016/0005-2787(72)90426-1. [DOI] [PubMed] [Google Scholar]
  2. Arnberg A. C., Van der Horst G., Tabak H. F. Formation of lariats and circles in self-splicing of the precursor to the large ribosomal RNA of yeast mitochondria. Cell. 1986 Jan 31;44(2):235–242. doi: 10.1016/0092-8674(86)90757-9. [DOI] [PubMed] [Google Scholar]
  3. Bruce A. G., Uhlenbeck O. C. Reactions at the termini of tRNA with T4 RNA ligase. Nucleic Acids Res. 1978 Oct;5(10):3665–3677. doi: 10.1093/nar/5.10.3665. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Cech T. R., Bass B. L. Biological catalysis by RNA. Annu Rev Biochem. 1986;55:599–629. doi: 10.1146/annurev.bi.55.070186.003123. [DOI] [PubMed] [Google Scholar]
  5. Fisher M. P., Dingman C. W. Role of molecular conformation in determining the electrophoretic properties of polynucleotides in agarose-acrylamide composite gels. Biochemistry. 1971 May 11;10(10):1895–1899. doi: 10.1021/bi00786a026. [DOI] [PubMed] [Google Scholar]
  6. Gough G. W., Lilley D. M. DNA bending induced by cruciform formation. Nature. 1985 Jan 10;313(5998):154–156. doi: 10.1038/313154a0. [DOI] [PubMed] [Google Scholar]
  7. Grabowski P. J., Zaug A. J., Cech T. R. The intervening sequence of the ribosomal RNA precursor is converted to a circular RNA in isolated nuclei of Tetrahymena. Cell. 1981 Feb;23(2):467–476. doi: 10.1016/0092-8674(81)90142-2. [DOI] [PubMed] [Google Scholar]
  8. Konarska M. M., Grabowski P. J., Padgett R. A., Sharp P. A. Characterization of the branch site in lariat RNAs produced by splicing of mRNA precursors. Nature. 1985 Feb 14;313(6003):552–557. doi: 10.1038/313552a0. [DOI] [PubMed] [Google Scholar]
  9. Loening U. E. The determination of the molecular weight of ribonucleic acid by polyacrylamide-gel electrophresis. The effects of changes in conformation. Biochem J. 1969 Jun;113(1):131–138. doi: 10.1042/bj1130131. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Marini J. C., Levene S. D., Crothers D. M., Englund P. T. Bent helical structure in kinetoplast DNA. Proc Natl Acad Sci U S A. 1982 Dec;79(24):7664–7668. doi: 10.1073/pnas.79.24.7664. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Peacock A. C., Dingman C. W. Resolution of multiple ribonucleic acid species by polyacrylamide gel electrophoresis. Biochemistry. 1967 Jun;6(6):1818–1827. doi: 10.1021/bi00858a033. [DOI] [PubMed] [Google Scholar]
  12. Reijnders L., Sloof P., Sival J., Borst P. Gel electrophoresis of RNA under denaturing conditions. Biochim Biophys Acta. 1973 Oct 26;324(3):320–333. doi: 10.1016/0005-2787(73)90278-5. [DOI] [PubMed] [Google Scholar]
  13. Ruskin B., Krainer A. R., Maniatis T., Green M. R. Excision of an intact intron as a novel lariat structure during pre-mRNA splicing in vitro. Cell. 1984 Aug;38(1):317–331. doi: 10.1016/0092-8674(84)90553-1. [DOI] [PubMed] [Google Scholar]
  14. Sugino A., Snoper T. J., Cozzarelli N. R. Bacteriophage T4 RNA ligase. Reaction intermediates and interaction of substrates. J Biol Chem. 1977 Mar 10;252(5):1732–1738. [PubMed] [Google Scholar]
  15. Tabak H. F., Van der Horst G., Kamps A. M., Arnberg A. C. Interlocked RNA circle formation by a self-splicing yeast mitochondrial group I intron. Cell. 1987 Jan 16;48(1):101–110. doi: 10.1016/0092-8674(87)90360-6. [DOI] [PubMed] [Google Scholar]
  16. Thorne H. V. Electrophoretic characterization and fractionation of polyoma virus DNA. J Mol Biol. 1967 Mar 14;24(2):203–211. doi: 10.1016/0022-2836(67)90326-9. [DOI] [PubMed] [Google Scholar]
  17. Thorne H. V. Electrophoretic separation of polyoma virus DNA from host cell DNA. Virology. 1966 Jun;29(2):234–239. doi: 10.1016/0042-6822(66)90029-8. [DOI] [PubMed] [Google Scholar]
  18. Wu H. M., Crothers D. M. The locus of sequence-directed and protein-induced DNA bending. Nature. 1984 Apr 5;308(5959):509–513. doi: 10.1038/308509a0. [DOI] [PubMed] [Google Scholar]
  19. van der Horst G., Tabak H. F. New RNA-mediated reactions by yeast mitochondrial group I introns. EMBO J. 1987 Jul;6(7):2139–2144. doi: 10.1002/j.1460-2075.1987.tb02481.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. van der Veen R., Arnberg A. C., van der Horst G., Bonen L., Tabak H. F., Grivell L. A. Excised group II introns in yeast mitochondria are lariats and can be formed by self-splicing in vitro. Cell. 1986 Jan 31;44(2):225–234. doi: 10.1016/0092-8674(86)90756-7. [DOI] [PubMed] [Google Scholar]

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