Analysis of gene function in Trypanosoma brucei using RNA interference.

Appolinaire Djikeng*, Shuiyuan Shen, Christian Tschudi, Elisabetta Ullu

*Corresponding author for this work

Research output: Contribution to journalArticlepeer-review

Abstract

Trypanosoma brucei, a flagellate protozoa of the family Trypanosomatidae, has become one of the model systems for unicellular pathogens to study fundamentally important biological phenomena. Currently, the method of choice to examine gene function in these organisms is RNA interference (RNAi). mRNA degradation is triggered by double-stranded RNA (dsRNA) produced in vivo from transgenes transcribed from opposing tetracycline (tet)-inducible T7 RNA polymerase promoters, or hairpin RNA transcribed from the tet-inducible procyclic acidic repetitive protein promoter. In this chapter, we describe some of the methods we employ for ablation of gene expression by RNAi in T. brucei with particular emphasis on transfection and cloning of procyclic cells, induction of dsRNA expression, isolation of RNA and analysis of dsRNA, and target mRNA.

Original languageEnglish
Pages (from-to)73-83
Number of pages11
JournalMethods in molecular biology (Clifton, N.J.)
Volume265
Publication statusPublished - 1 Jan 2004

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