Abstract
BackgroundRNA interference (RNAi) is a powerful tool to study gene function in organisms that are not amenable to classical forward genetics. Hence, together with the ease of comprehensively identifying genes by new generation sequencing, RNAi is expanding the scope of animal species and questions that can be addressed in terms of gene function. In the case of genetic mutants, the genetic background of the strains used is known to influence the phenotype while this has not been described for RNAi experiments.ResultsHere we show in the red flour beetle Tribolium castaneum that RNAi against Tc-importin α1 leads to different phenotypes depending on the injected strain. We rule out off target effects and show that sequence divergence does not account for this difference. By quantitatively comparing phenotypes elicited by RNAi knockdown of four different genes we show that there is no general difference in RNAi sensitivity between these strains. Finally, we show that in case of Tc-importin α1 the difference depends on the maternal genotype.ConclusionsThese results show that in RNAi experiments strain specific differences have to be considered and that a proper documentation of the injected strain is required. This is especially important for the increasing number of emerging model organisms that are being functionally investigated using RNAi. In addition, our work shows that RNAi is suitable to systematically identify the differences in the gene regulatory networks present in populations of the same species, which will allow novel insights into the evolution of animal diversity.
Highlights
RNA interference (RNAi) is a powerful tool to study gene function in organisms that are not amenable to classical forward genetics
Phylogenetic analysis revealed that the TC000963 gene is the single importin α1 ortholog of Tribolium, called Tc-importin α1 in the following (Figure 1, red frame; see Additional file 1 C for aligned sequences)
The San Bernadino (SB) typical “abdomen inside-out” phenotype was not observed. These results show that it is primarily the genotype of the mother, which determines the quality of the Tc-importin α1 RNAi phenotype while the minor increase of “labrum only” phenotypes to 4,3% indicate some influence of the zygotic genome, too
Summary
RNA interference (RNAi) is a powerful tool to study gene function in organisms that are not amenable to classical forward genetics. In the case of genetic mutants, the genetic background of the strains used is known to influence the phenotype while this has not been described for RNAi experiments. The identification of gene functions has been based on classical forward genetic screens where mutants are randomly generated, e.g. by chemical or transposon mediated mutagenesis. The established mutant strains are screened for phenotypes and subsequently the disrupted gene is identified and further analyzed [1,2,3,4,5]. The unbiased forward genetic approach to identify gene functions has been very successful but it limits the questions that can be
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