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

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.

Sublethal effects of chlorantraniliprole on juvenile hormone levels and mRNA expression of JHAMT and FPPS genes in the rice stem borer, Chilo suppressalis

“…The procedures for RT‐qPCR were the same as those described by Zhu et al . The elongation factor 1 ( EF‐1 ) gene was used as an internal control . The PCR reaction volume was 20 μL containing 2 μL of diluted cDNA, 0.4 µ m of each primer, 10 μL SYBR Premix EX Taq™ II (2×) and 0.4 μL ROX Reference Dye II (50×).…”

“…The procedures for RT-qPCR were the same as those described by Zhu et al 34 The elongation factor 1 (EF-1) gene was used as an internal control. 35,36 The PCR reaction volume was 20 L containing 2 L of diluted cDNA, 0.4 μM of each primer, 10 L SYBR Premix EX Taq™ II (2×) and 0.4 L ROX Reference Dye II (50×). PCR conditions were set as an initial incubation of 95 ∘ C for 30 s; 40 cycles of 95 ∘ C for 5 s and 60 ∘ C for 30 s. After amplification, a melting curve analysis was performed to confirm the specificity of the RT-qPCR reaction and lack of primer dimers.…”

Downregulation of Chilo suppressalis alkaline phosphatase genes associated with resistance to three transgenic Bacillus thuringiensis rice lines

“…Total RNA was extracted from the whole bodies of 15 larvae from each dsRNA treatment group with three replicate samples for each treatment group. qRT-PCR primers were designed using the National Center for Biotechnology Information profile server (http://www.ncbi.nlm.nih.gov/tools/primer-blast; Supporting Information Table S3) with the C. suppressalis elongation factor-1 gene as the internal reference (Zhu et al, 2016;Qiu et al, 2017). The efficiency of the qRT-PCR primers used was determined (Supporting Information Table S3) using the protocol described in Qiu et al (2017).…”

The midgut V‐ATPase subunit A gene is associated with toxicity to crystal 2Aa and crystal 1Ca‐expressing transgenic rice in Chilo suppressalis

“…(). The C. suppressalis elongation factor‐1 gene (Table ) was used as the internal reference (Zhu et al ., ; Qiu et al ., ). The qRT‐PCR protocol was similar to that described previously (Qiu et al ., ).…”