A doubly inducible system for RNA interference and rapid RNAi plasmid construction in Trypanosoma brucei

摘要: The most rapid method for the generation of conditional mutants in Trypanosoma brucei is use RNA interference. A single copy target sequence cloned between two opposing T7 promoters bearing tet operators, and resulting plasmid integrated into genome cells expressing both repressor polymerase. Upon addition tetracycline, double-stranded synthesised from promoters. Unfortunately, repression promoter activity may sometimes be insufficient to prevent expression toxic amounts RNA. We describe here cell lines which polymerase under tetracycline control, show that regulation can modulate transcription a constitutive promoter. In we construct containing copies tn10 Tet easy creation repressor-expressing trypanosomes, an interference vector allows "TA" cloning unmodified PCR products blue/white selection.