Inducible RNAi as a forward genetic tool in Trypanosoma brucei

Mark E. Drew, Shawn A. Motyka, James C. Morris, Zefeng Wang, Paul T. Englund

Research output: Chapter in Book/Report/Conference proceedingChapter

Abstract

Introduction to RNAi in Trypanosomes In December 1998 the Ullu lab at Yale published the first report of dsRNA-mediated mRNA degradation in Trypanosoma brucei (Ngo et al., 1998). These experiments used either electroporation of in vitro synthesized dsRNA or transient in vivo expression of single-strand RNA that forms a stem–loop structure capable of inducing RNAi. At that time, the arsenal of genetic techniques available to the trypanosome researcher was limited (Clayton, 1999). For example, gene knockout was possible through homologous recombination, although the diploid genome of T. brucei complicated this approach. Furthermore, essential genes were difficult to examine by knockout, necessitating complex strategies in which inducible ectopic expression needed to be maintained while the genomic knockouts were generated (Wirtz et al., 1999). RNAi raised hopes for a powerful and convenient genetic approach for these eukaryotes. Our lab has made extensive use of RNAi in studying gene function in T. brucei, the eukaryotic parasite that causes African sleeping sickness. The purpose of this chapter is to review the steps our lab has taken in developing an inducible RNAi system that has allowed us to achieve the goal of bona fide RNAi-based forward genetics in T. brucei. In addition, this chapter will review our development of an easy-to-use, inducible RNAi system, presenting a few examples of how this approach has allowed us to gain new insights into gene function, especially in the case of essential genes.

Original languageEnglish (US)
Title of host publicationRNA Interference Technology: From Basic Science to Drug Development
PublisherCambridge University Press
Pages247-256
Number of pages10
ISBN (Print)9780511546402, 0521836778, 9780521836777
DOIs
StatePublished - Jan 1 2005

Fingerprint

Trypanosoma brucei brucei
RNA Interference
Genes
Trypanosomiasis
Essential Genes
Arsenals
African Trypanosomiasis
Genetic Techniques
Gene Knockout Techniques
Electroporation
Homologous Recombination
RNA Stability
Eukaryota
Diploidy
RNA
Parasites
Degradation
Research Personnel
Messenger RNA
Genome

ASJC Scopus subject areas

  • Biochemistry, Genetics and Molecular Biology(all)

Cite this

Drew, M. E., Motyka, S. A., Morris, J. C., Wang, Z., & Englund, P. T. (2005). Inducible RNAi as a forward genetic tool in Trypanosoma brucei. In RNA Interference Technology: From Basic Science to Drug Development (pp. 247-256). Cambridge University Press. https://doi.org/10.1017/CBO9780511546402.021

Inducible RNAi as a forward genetic tool in Trypanosoma brucei. / Drew, Mark E.; Motyka, Shawn A.; Morris, James C.; Wang, Zefeng; Englund, Paul T.

RNA Interference Technology: From Basic Science to Drug Development. Cambridge University Press, 2005. p. 247-256.

Research output: Chapter in Book/Report/Conference proceedingChapter

Drew, ME, Motyka, SA, Morris, JC, Wang, Z & Englund, PT 2005, Inducible RNAi as a forward genetic tool in Trypanosoma brucei. in RNA Interference Technology: From Basic Science to Drug Development. Cambridge University Press, pp. 247-256. https://doi.org/10.1017/CBO9780511546402.021
Drew ME, Motyka SA, Morris JC, Wang Z, Englund PT. Inducible RNAi as a forward genetic tool in Trypanosoma brucei. In RNA Interference Technology: From Basic Science to Drug Development. Cambridge University Press. 2005. p. 247-256 https://doi.org/10.1017/CBO9780511546402.021
Drew, Mark E. ; Motyka, Shawn A. ; Morris, James C. ; Wang, Zefeng ; Englund, Paul T. / Inducible RNAi as a forward genetic tool in Trypanosoma brucei. RNA Interference Technology: From Basic Science to Drug Development. Cambridge University Press, 2005. pp. 247-256
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