The piggyBac mobile element is quickly gaining popularity as a tool for the transgenesis of many eukaryotic organisms. By studying the transposase (TPase) which catalyzes the movement of piggyBac, modification of this vector system may be possible in order to make it a more effective transgenesis tool. In a previous publication (Sarkar et al., 2003) proposed the presence of the widespread 'DDE/DDD' motif for piggyBac at amino acid positions D268, D346, and D447. A PSORTII analysis of the TPase amino acid sequence predicts several nuclear localization signals (NLS) near the C-terminus, just upstream of a putative zinc finger (ZnF). This study utilizes directed mutagenesis and plasmid-based mobility assays to assess the importance of conserved aspartate residues as the catalytic core of the piggyBac TPase. Individual point-mutations have been functionally analyzed with respect to charge and physical size in all three proposed residues of the 'DDD' motif, as well as another nearby, highly conserved aspartate at D450. Results indicate that all four aspartates are necessary, to one degree or another, for excision to occur in a cellular environment, but D450 seems to have a tolerance for a glutamate substitution. The piggyBac TPase was fused upstream and in-frame with the enhanced yellow fluorescent protein (EYFP) in the Drosophila melanogaster inducible metallothionein (MT) promoter and the fluorescence tracked by confocal microscopy. Through N and C-terminal truncations, targeted internal deletions, and specific amino acid mutations of the piggyBac TPase open reading frame (ORF), the region containing the PSORTII predicted NLSs was determined to be required for the TPase to enter the nucleus of S2 cells; other additional negatively charged amino acids a short length upstream of this region were also required for proper function of the NLS. Control of the spread of dengue virus (DENV) has become a more important concern, with the growth of worldwide travel and development of rural areas. Recent outbreaks in the third world have taken a heavy economic toll on developing countries in terms of morbidity. An effective vaccine against all four serotypes of the virus has remained elusive due to antibody dependent enhancement of a viral infection. One alternative to vaccines currently being explored is the creation of mosquitoes unable to spread the virus from person to person. The establishment and spread of a so called refractory strain would effectively halt the DENV life cycle in its tracks. Obviously, a key element of this population replacement approach is an effective transgene conferring resistance to viral transmission. The ability to precisely target the transgene to invariant elements of the virus would make it more effective against a wider variety of serotypes and strains. The ability of the Tetrahymena thermophila group I intron to catalyze the trans-splicing of subgenomic sequences highly conserved in the dengue genome across all serotypes was tested. By specifically targeting these regions, trans-splicing products were detected both in vitro with T7 RNA transcripts and in situ from traditional RNA polymerase II (RNAP II) promoted transcripts. A number of different introns, which splice with varying efficiencies as measured by real-time reverse transcription polymerase chain reaction (PCR) (qRT-PCR) and Dual Luciferase assay, have been designed and tested. The trans-splicing of the group I intron can be directed to splice a novel 3' exon to the upstream fragment of the splicing product, allowing for de novo gene expression upon detection of the target. When linked to a pro-apoptotic gene as a 3' exon and expressed constitutively in a cell, the group I intron may trigger ordered cell death upon infection, denying the virus the ability to reproduce. One such pro-apoptotic gene, the truncated bcl-2-associated protein X (tBax), was tested for its ability to induce apoptosis in insect cells and found to be an effective pro-apoptotic gene.