ARS | Publication request: Illumina sequencing of green stink bug nymph and adult cdna to identify potential rnai gene targets

Submitted to: Meeting Proceedings
Publication Type: Proceedings
Publication Acceptance Date: January 18, 2012
Publication Date: July 25, 2012
Citation: Van Kretschmar, J.B., Donohue, K.V., Cabrera Cordon, A.R., Magalhaes, L.C., Sorenson, C.E., Bacheler, J.S., Khalil, S.M., Roe, R.M. 2012. Illumina sequencing of green stink bug nymph and adult cdna to identify potential rnai gene targets. Beltwide Cotton Conferences. 1090-1096.

Technical Abstract: Whole-body transcriptomes for nymphs and adults of the green stink bug, Acrosternum hilare (Say), were sequenced on an Illumina® Genome Analyzer IIx sequencer. The insects were collected from sites in North Carolina and Virginia, USA. The cDNA library for each sample was sequenced on one lane of an eight-lane flow cell. Total read count was 25,211,411 and 23,601,951 for the nymph and adult libraries, respectively. Mean read length was 124 bps (base pairs) for nymph and 125 bps for adult. Total sequencing data were 3,127,036,121 bps for nymph cDNA and 2,956,794,438 bps for adult. Trinity® software was used to assemble the nymph and adult reads into 79,062 and 74,271 contigs (contiguous sequences), respectively. Blast2GO® software was used to align, map, and annotate the contigs. For the alignment step, the contigs were translated to peptides and compared to the GenBank nr (non-redundant) protein database using the BLASTx (Basic Local Alignment Search Tool) algorithm with E-value cut-off set at E-3 (10-3). The average length of BLASTx translated sequences was 1248 (range = 201 – 7968) amino acids for the nymph and 1320 (range = 201 – 7957) for the adult. Several nymph and adult sequences showed high homology (E-value = 1.5 E-13) with database hormones, neuropeptides, neurotransmitters and/or their receptors. Seventeen nymph and adult sequences were identical (E-value = 0) to A. pisum, T. castaneum, B. germaninca, and P. humanus corporis hormones. After alignment of green stink bug contigs with GenBank nr database sequences, all BLAST hits (E-value = E-3) were mapped and annotated with GO (Gene Ontology) terms that assigned the translated sequences to categories of putative protein function. Mapping and annotation steps yielded 13,226 nymph and 14,725 adult sequences assigned to 13 GO functional categories: Catalysis, binding, transport, signal transduction, structural molecule activity, enzyme regulation, electron carrier activity, antioxidant activity, metallochaperone, channel regulation, protein tag, translation regulation and nutrient reservoir. The goal of sequencing and of on-going bioinformatic analysis of these transcriptomes is to identify candidate RNAi gene targets with potential to be developed as insecticidal transgenes for control of stink bugs and other hemipteran pests of cotton. The resulting data can also be applied to other protein function and genetics studies.