The hAT -family transposable element, hopper , from Bactrocera dorsalis is a functional vector for insect germline trans
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RESEARCH
Open Access
The hAT-family transposable element, hopper, from Bactrocera dorsalis is a functional vector for insect germline transformation Alfred M. Handler1* and Marc F. Schetelig2
Abstract Background: The hopper hAT-family transposable element isolated from the Oriental fruit fly, Bactrocera dorsalis, is distantly related to both the Drosophila hobo element and the Activator element from maize. The original 3120 bp hopperBd-Kah element isolated from the Kahuku wild-type strain was highly degenerate and appeared to have a mutated transposase and terminal sequences, while a second 3131 bp element, hopperBd-we, isolated from a white eye mutant strain had an intact transposase reading frame and terminal sequences consistent with function. Results: The hopperBd-we element was tested for function by its ability to mediate germline transformation in two dipteran species other than B. dorsalis. This was achieved by creating a binary vector/helper transformation system by linking the hopperBd-we transposase reading frame to a D. melanogaster hsp70 promoter for a heat-inducible transposase helper plasmid, and creating vectors marked with the D. melanogaster mini-white+ or polyubiquitinregulated DsRed fluorescent protein markers. Conclusions: Both vectors were successfully used to transform D. melanogaster, and the DsRed vector was also used to transform the Caribbean fruit fly, Anastrepha suspensa, indicating a wide range of hopper function in dipteran species and, potentially, non-dipteran species. This vector provides a new tool for insect genetic modification for both functional genomic analysis and the control of insect populations. Keywords: Insect genetic modification, Transposon-mediated transformation, Tephritidae
Background Transposon-mediated germline transformation has been the primary method of insect genomic manipulation since a P element vector was successfully transposed into the Drosophila melanogaster genome [1]. More recent gene-editing techniques, such as CRISPR/Cas9, have provided additional methods for genome manipulation, but thus far are limited in achieving genomic * Correspondence: [email protected] 1 USDA/ARS, Center for Medical, Agricultural and Veterinary Entomology, 1700 SW 23rd Drive, Gainesville, FL 32608, USA Full list of author information is available at the end of the article
integration of DNA constructs greater than several kilobases [2]. This limitation is especially critical for the development of genetically modified strains to improve biologically-based strategies for the control of insect populations harmful to agriculture and human health, for which conditional lethal and other transgene constructs are typically 10 kb or greater. Significantly, geneediting is also limited in generating random genomic insertions for mutagenesis and enhancer-trap screens that have proven highly advantageous for functional genomic analysis, most clearly demonstrated by elucidating gene function and regulation in the D. melanogaster model
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