Multi-toxin Bt-crops expressing insecticidal toxins with different modes of action e.g. Cry and Vip, are expected to improve resistance management in target pests. While Cry1A resistance has been relatively well characterized in some insect species, this is not the case for Vip3A, for which no mechanism of resistance has yet been identified. Here we applied HT-SuperSAGE to analyze the transcriptome of the gut tissue of tobacco budworm Heliothis virescens (F.) laboratory-selected for Vip3Aa resistance. From a total of 1 324 252 sequence reads, 58951 26-bp tags were obtained representing 17 751 non-singleton unique transcripts (UniTags) from genetically similar Vip3Aa-resistant (Vip-Sel) and susceptible control (Vip-Unsel) strains. Differential expression was significant (≥ 2.5-fold or ≤ 0.4; P<0.05) for 1989 sequences (11.2% of total UniTags), where 420 represented over-expressed (OE) and 1569 under-expressed (UE) genes in Vip-Sel. BLASTN searches mapped 419 UniTags to H. virescens sequence contigs, of which, 416 (106 OE and 310 UE) were unambiguously annotated to proteins in NCBI non-redundant protein databases. Gene Ontology distributed 345 of annotated UniTags in 14 functional categories with metabolism (including serine-type hydrolases) and translation/ribosome biogenesis being the most prevalent. A UniTag homologous to a particular member of the REsponse to PAThogen (REPAT) family was found among most over-expressed, while, UniTags related to the putative Vip3Aa-binding ribosomal protein S2 (RpS2) were under-expressed. qRT-PCR of a subset of UniTags validated the HT-SuperSAGE data. The present study is the first providing lepidopteran gut transcriptome associated with Vip3Aa resistance and a foundation for future attempts to elucidate the resistance mechanism. This article is protected by copyright. All rights reserved.