Promoter and piggyBac activities within embryos of the potato tuber moth, Phthorimaea operculella, Zeller (Lepidoptera: Gelechiidae).
Potato production in tropical and subtropical countries suffers from damage caused by the potato tuber moth (PTM), Phthorimaea operculella. The aim of this research was the development of the components required for a germline transformation system for the PTM. We tested three components that are critical to genetic transformation systems for insects: promoter activity, marker gene expression, and transposable element function. We compared the transcriptional activities of five different promoters, hsp70, hsp82, actin5C, polyubiquitin and immediate early 1 gene (ie1), within PTM embryos. The ie1 promoter, flanked by the hr5 enhancer element, showed a very high level of transcriptional activity compared to the other promoters. The fluorescence activity of EGFP was also determined and transient expression of EGFP was detected in 57% of injected embryos. The transpositional activity of the piggyBac transposable element was tested in an interplasmid transposition assay. The piggyBac element was shown to be mobile within the embryonic soma of the PTM with a transposition frequency of 4.2 x 10(-5) transpositions/donor plasmid. Incorporating a transactivator plasmid expressing the immediate early protein (IE1) from the Bombyx mori nuclear polyhedrosis virus enhanced the efficiency of piggyBac mobility.
Mohammed
Ahmed
A
Department of Entomology, Texas A&M University, 2475 TAMU, College Station, TX 77843-2475, USA.
Coates
Craig J
CJ
D000199:Actins / Q000235:genetics; D000818:Animals; D004251:DNA Transposable Elements / Q000235:genetics*; D004625:Embryo, Nonmammalian / Q000378:metabolism*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D018840:HSP70 Heat-Shock Proteins / Q000235:genetics; D018841:HSP90 Heat-Shock Proteins; D006360:Heat-Shock Proteins / Q000235:genetics; D017874:Immediate-Early Proteins / Q000235:genetics; D008156:Luciferases / Q000235:genetics / Q000378:metabolism; D008845:Microinjections; D009036:Moths / Q000196:embryology / Q000235:genetics*; D016254:Mutagenesis, Insertional; D010957:Plasmids / Q000008:administration & dosage / Q000235:genetics; D025821:Polyubiquitin / Q000235:genetics; D011401:Promoter Regions, Genetic / Q000235:genetics*; D011993:Recombinant Fusion Proteins / Q000235:genetics / Q000378:metabolism; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators / Q000235:genetics
D000199:Actins; D004251:DNA Transposable Elements; D018840:HSP70 Heat-Shock Proteins; C074510:HSP82 protein, S cerevisiae; D018841:HSP90 Heat-Shock Proteins; D006360:Heat-Shock Proteins; D017874:Immediate-Early Proteins; D011993:Recombinant Fusion Proteins; D029701:Saccharomyces cerevisiae Proteins; D015534:Trans-Activators; D025821:Polyubiquitin; D049452:Green Fluorescent Proteins; D008156:Luciferases