Larval mouthparts from Helicoverpa assulta
Total RNA was purified using RNeasy (Qiagen, USA) kit according to the manufacturer's protocol. To remove the contaminated genomic DNA, the purified RNA was treated with DNaseI (Ambion, USA), then quantified and qualified by NanoDrop ND-2000 (Thermo Scientific, USA) as well as 2100 Bioanalyzer (Agilent, USA). Prepared RNA samples from larval antennae, larval mouthparts, male adult head and male adult tarsi were sent to Beijing Genomic Institute (BGI, http://www.genomics.cn/index.php) for the generation of Paired-End Illumina TruSeq libraries and sequencing with an Illumina HiSeq 2000 platform. cDNA templates were prepared from purified RNA samples of male and female adult antennae using SuperScript III Reverse Transcriptase (Invitrogen, USA), according to the manufacturer's manual. RT-PCR was performed using gene-specific primers (GSPs) as follows: 95 C for 3 min; 35 cycles at 95 C for 30 s, 50 C for 30 s and 72 C for 45 s; and final extension at 72 C for 10 min. PCR products were analysed by using 1.5% agarose gels. HassOR2 (Orco or Or83b) gene was used as control to justify the quality of synthesized cDNA samples (Xu et al., 2010). Each RT-PCR reaction was repeated at least 3 times.