Preview

Fine Chemical Technologies

Advanced search

Test system creation for analysis of template switching effect during a reverse transcription in vitro

Abstract

We investigated an in vitro system for the analysis of template switching effect of RNA-dependent DNA-polymerase during reverse transcription reaction. An RT-PCR-based approach was realized. An RNA sample was used as a template. The RNA contains a region with a stable secondary structure flanked by two homologous sequences. The described system can find use for testing overall performance of new and existing forms of the reverse transcriptase.

About the Authors

A. Y. Komkov
Институт биоорганической химии им. академиков М.М. Шемякина и Ю.А. Овчинникова РАН
Russian Federation


Y. B. Lebedev
Институт биоорганической химии им. академиков М.М. Шемякина и Ю.А. Овчинникова РАН
Russian Federation


V. I. Shvets
M.V. Lomonosov Moscow State University of Fine Chemical Technologies, 86, Vernadskogo pr., Moscow 119571
Russian Federation


References

1. Amariglio N., Rechavi G. A-to-I RNA editing: А new regulatory mechanism of global gene expression // Blood Cells Mol. Dis. 2007. V. 39. № 2. p. 151-155.

2. Maas S., Kawahara Y., Tamburro K.M., Nishikura K. A-to-I RNA editing and human disease // RNA Biol. 2006. V. 3. № 1. p. 1-9.

3. Lopez-Saavedra A., Herrera L.A. The role of alternative mRNA splicing in chromosome instability // Mutat. Res. 2010. V. 705. № 3. p. 246-251.

4. Hallegger M., Llorian M., Smith C.W. Alternative splicing: Global insights // FEBS J. 2010. V. 277. № 4. p. 856-866.

5. David C. J., Manley J.L. Alternative pre-mRNA splicing regulation in cancer: Pathways and programs unhinged // Genes Dev. 2010. V. 24. № 21. p. 2343-2364.

6. Khanna A., Stamm S. Regulation of alternative splicing by short non-coding nuclear RNAs // RNA Biol. 2010. V. 7. № 4. p. 480-485.

7. Huang Y., Shen X.J., Zou Q., Zhao Q.L. Biological functions of microRNAs // Биоорган. химия. 2010. Т. 36. № 6. С. 747-752.

8. Huntzinger E., Izaurralde E. Gene silencing by microRNAs: Сontributions of translational repression and mRNA decay // Nat. Rev. Genet. 2011. V. 12. № 2. p. 99-110.

9. Wang K.C., Chang H.Y. Molecular mechanisms of long noncoding RNAs // Mol. Cell. 2011. V. 43. № 6. p. 904-914.

10. Wapinski O., Chang H.Y. Long noncoding RNAs and human disease // Trends Cell Biol. 2011. V. 21. № 6. p. 354-361.

11. Liao Q., Liu C., Yuan X., Kang S., Miao R., Xiao H., Zhao G., Luo H., Bu D., Zhao H., Skogerbо G., Wu Z., Zhao Y. Large-scale prediction of long non-coding RNA functions in a coding-non-coding gene co-expression network // Nucleic Acids Res. 2011. V. 39. № 9. p. 3864-3878.

12. Temin H.M. Proceedings: Molecular mechanisms for cancer induction by viruses // Proc. Natl. Cancer Conf. 1972. V. 7. P. 415-419.

13. Gilboa E., Mitra S. W., Goff S., Baltimore D. A detailed model of reverse transcription and tests of crucial aspects // Cell. 1979. V. 18. № 1. p. 93-100.

14. Temin H.M. Retrovirus variation and reverse transcription: abnormal strand transfers result in retrovirus genetic variation // Proc. Natl. Acad. Sci. USA. 1993. V. 90. № 15. p. 6900-6903.

15. Roy S. W., Irimia M. When good transcripts go bad: Artifactual RT-PCR 'splicing' and genome analysis // Bioessays. 2008. V. 30. № 6. p. 601-605.

16. Cocquet J., Chong A., Zhang G., Veitia R.A. Reverse transcriptase template switching and false alternative transcripts // Genomics. 2006. V. 88. № 1. p. 127-131.

17. Houseley J., Tollervey D. Apparent non-canonical trans-splicing is generated by reverse transcriptase in vitro // PLoS One. 2010. V. 5. № 8. p. e12271.

18. International human genome consortium. Initial sequencing and analysis of the human genome // Nature. 2001. V. 409. № 6822. p. 860-921.

19. Osenberg S, Dominissini D, Rechavi G., Eisenberg E. Widespread a cleavage of A-to-I hyperediting substrates // RNA. 2009. V. 15. № 9. P. 1632-1639.

20. Sambrook J., Russel D. Molecular cloning: A laboratory manual. - New York: Cold Spring Harbor Laboratory Press, 2001. 2344 p.


Review

For citations:


Komkov A.Y., Lebedev Y.B., Shvets V.I. Test system creation for analysis of template switching effect during a reverse transcription in vitro. Fine Chemical Technologies. 2011;6(6):77-83. (In Russ.)

Views: 332


ISSN 2410-6593 (Print)
ISSN 2686-7575 (Online)