The effect of silencing 20E biosynthesis relative genes by feeding bacterially expressed dsRNA on the larval development of Chilo suppressalis.

Jian Zhu,Ping Li,Chang-Ying Niu,Yong-Cheng Dong

doi:10.1038/srep28697

Jian Zhu, Ping Li + Show 2 more

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https://doi.org/10.1038/srep28697

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Journal: Scientific Reports	Publication Date: Jun 29, 2016
Citations: 26	License type: CC BY 4.0

Affiliation: Huazhong Agricultural University

Abstract

RNA interference (RNAi) is a robust tool to study gene functions as well as potential for insect pest control. Finding suitable target genes is the key step in the development of an efficient RNAi-mediated pest control technique. Based on the transcriptome of Chilo suppressalis, 24 unigenes which putatively associated with insect hormone biosynthesis were identified. Amongst these, four genes involved in ecdysteroidogenesis i.e., ptth, torso, spook and nm-g were evaluated as candidate targets for function study. The partial cDNA of these four genes were cloned and their bacterially expressed dsRNA were fed to the insects. Results revealed a significant reduction in mRNA abundance of target genes after 3 days. Furthermore, knocked down of these four genes resulted in abnormal phenotypes and high larval mortality. After 15 days, the survival rates of insects in dsspook, dsptth, dstorso, and dsnm-g groups were significantly reduced by 32%, 38%, 56%, and 67% respectively, compared with control. Moreover, about 80% of surviving larvae showed retarded development in dsRNA-treated groups. These results suggest that oral ingestion of bacterially expressed dsRNA in C. suppressalis could silence ptth, torso, spook and nm-g. Oral delivery of bacterially expressed dsRNA provides a simple and potential management scheme against C. suppressalis.

Highlights

All unigene sequences were annotated by searching the Nr (NCBI non-redundant protein sequences) protein database using BLASTx with an E-value threshold of 10−5 (1.0E-5), a total number of 19,990 distinct sequences (43% of all unigenes) were matched with known genes that encoded for functional proteins
For example in Helicoverpa armigera, the brain is the major site for ptth expression and low levels of expression were reported in the midgut and fat body36. ptth has been detected in different tissues of Sesamia nonagrioides, including brain, ganglion, gut and fat body[37], the function of prothoracicotropic hormone (PTTH) produced in other organs still remains unclear
The transcriptome of C. suppressalis was sequenced with Illumina that yielded 46,603 unigenes

Summary

Introduction

All unigene sequences were annotated by searching the Nr (NCBI non-redundant protein sequences) protein database using BLASTx with an E-value threshold of 10−5 (1.0E-5), a total number of 19,990 distinct sequences (43% of all unigenes) were matched with known genes that encoded for functional proteins. Based on E-value distribution of top hits, 60% of the mapped unigene sequences showed significant homology (less than 1.0E-50) with deposited genes in Nr database (Fig. 1A). By searching against the same database, 73% and 33% of the unigenes were found to possess sequence similarities greater than 50% and 75%, respectively (Fig. 1B). With respect to organism sources of homologs, almost 98% of unigenes with Nr annotations were matched to animal genes (Fig. 1C). While 58%, 8%, 5%, 3%, 3% of the mapped unigene sequences possess significant similarity, with orthologs Danaus plexippus, Bombyx mori, Tribolium castaneum, Papilio xuthus and Acyrthosiphon pisum respectively (Fig. 1D)

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