Identifying potential RNAi targets in grain aphid (Sitobion avenae F.) based on transcriptome profiling of its alimentary canal after feeding on wheat plants

Abstract

BackgroundThe grain aphid (Sitobion avenae F.) is a major agricultural pest which causes significant yield losses of wheat in China, Europe and North America annually. Transcriptome profiling of the grain aphid alimentary canal after feeding on wheat plants could provide comprehensive gene expression information involved in feeding, ingestion and digestion. Furthermore, selection of aphid-specific RNAi target genes would be essential for utilizing a plant-mediated RNAi strategy to control aphids via a non-toxic mode of action. However, due to the tiny size of the alimentary canal and lack of genomic information on grain aphid as a whole, selection of the RNAi targets is a challenging task that as far as we are aware, has never been documented previously.ResultsIn this study, we performed de novo transcriptome assembly and gene expression analyses of the alimentary canals of grain aphids before and after feeding on wheat plants using Illumina RNA sequencing. The transcriptome profiling generated 30,427 unigenes with an average length of 664 bp. Furthermore, comparison of the transcriptomes of alimentary canals of pre- and post feeding grain aphids indicated that 5490 unigenes were differentially expressed, among which, diverse genes and/or pathways were identified and annotated. Based on the RPKM values of these unigenes, 16 of them that were significantly up or down-regulated upon feeding were selected for dsRNA artificial feeding assay. Of these, 5 unigenes led to higher mortality and developmental stunting in an artificial feeding assay due to the down-regulation of the target gene expression. Finally, by adding fluorescently labelled dsRNA into the artificial diet, the spread of fluorescence signal in the whole body tissues of grain aphid was observed.ConclusionsComparison of the transcriptome profiles of the alimentary canals of pre- and post-feeding grain aphids on wheat plants provided comprehensive gene expression information that could facilitate our understanding of the molecular mechanisms underlying feeding, ingestion and digestion. Furthermore, five novel and effective potential RNAi target genes were identified in grain aphid for the first time. This finding would provide a fundamental basis for aphid control in wheat through plant mediated RNAi strategy.

Identifying potential RNAi targets in aphid species (Macrosiphum rosae, Rhopalosiphum maidis, Sitobion avenae, Toxoptera aurantii) in sub-tropical region with extreme summer

“…Based on this observation 5 major exotic unigenes explored via transcriptomic profiling of wheat aphids were selected (Zhang et al, 2013). These unigenes were confirmed in our local aphid strains such as M. rosae, R. maidis, S. avenae, T. aurantii) feeding on C. sinensis, R. indica, S. melongen, Z. mays and T. aestivum.…”

“…Secondly artificial feed also because high mortality within 24 hours comparable to microinjection and low dosage of the construct in artificial diet was found to be efficient for aphid mortality. The 5 unigene sequences selected will be used for RNAi based assay by producing transgenic tobacco followed by transgenic corn and wheat (Zhang et al, 2013).…”

“…30,427 novel unigenes were identified in grain aphids as successful target of RNAi after gene expression analysis of their alimentary canal. From all these unigenes, 5 were reported to favor minimal risk of non-specific target but also maintain the low efficacy of lethal dose in comparison to previous studies (Zhang et al, 2013;Pitino et al, 2011). These target genes after their confirmation and introduction in our local wheat varieties will be efficient for aphid control in local agricultural practice.…”

“…Sequence of five unigenes provided by Zhang et al (2013) were used to design relatively suitable length of PCR primers using primer 3 online software (version 0.4.0; Rozen and Skaletsky, 2000).…”

Wheat Biotechnology: Current Status and Future Prospects

“…It has been demonstrated that a range of insect cells possess RNAi activity (Roether and Meister 2011;Terenius et al 2011). With the purpose of identifying target genes for silencing in aphid gut, Zhang et al (2013) performed RNA sequencing on the alimentary canals of grain aphids before and after feeding on wheat plants. They identified 16 genes that were significantly up or downregulated upon feeding and suggested these were good targets for RNAi insect control strategies.…”