Host-delivered RNAi mediated silencing of HaAce1, encoding the major isoform of acetylcholinesterase, imparts resistance against the polyphagous insect-pest, Helicoverpa armigera

Abstract The polyphagous insect pest, Helicoverpa armigera, is a detriment to crop productivity. Host-delivered RNAi has emerged as a potential tool to complement Bt technology for controlling insect pest menace. In this study transgenic tobacco ( Nicotiana tabacum L.; cv. Petit Havana) plants expressing dsRNA targeting the H. armigera gene, HaAce1 , encoding the major isoform of acetylcholinesterase, were developed. A 643 nucleotide RT-PCR amplified HaAce1 cDNA fragment was ligated in sense and antisense orientation intervened by GBSS intron to develop an inverted repeat (IR) gene construct. The HaAce1 IR gene construct (IR- Ace1 ) under the transcriptional control of CaMV 35S promoter and NOS terminator was used for Agrobacterium -mediated transformation of tobacco leaf disc explants. Fourteen HaAce1- hpRNA tobacco transgenic lines were obtained after screening of 31 putative transformants by PCR and RT-PCR. Five HaAce1- hpRNA tobacco transgenic lines demonstrated high level of resistance against H. armigera larvae based on detached leaf insect bioassay. These selected five tobacco transgenic lines carried two to four copies of the transgene . Generation of HaAce1 -specific siRNA was detected in these tobacco transgenic lines. Semi-quantitative PCR revealed many-fold reductions in the steady-state level of HaAce1 mRNA in larvae fed on leaves of the selected transgenic tobacco lines compared to that in control tobacco leaf-fed larvae. Our findings demonstrate that the host delivered dsRNA targeting Ace1 gene could be a promising strategy for controlling insect pest menace in crops.