Mariner transposition and transformation of the yellow fever mosquito, Aedes aegypti

Abstract

The mariner transposable element is capable of interplasmid transposition in the embryonic soma of the yellow fever mosquito, Aedes aegypti. To determine if this demonstrated mobility could be utilized to genetically transform the mosquito, a modified mariner element marked with a wild-type allele of the Drosophila melanogaster cinnabar gene was microinjected into embryos of a kynurenine hydroxylasedeficient, white-eyed recipient strain. Three of 69 fertile male founders resulting from the microinjected embryos produced families with colored-eyed progeny individuals, a transformation rate of 4%. The transgene-mediated complementation of eye color was observed to segregate in a Mendelian manner, although one insertion segregates with the recessive allele (female-determining) of the sex-determining locus, and a separate insertion is homozygous lethal. Molecular analysis of selected transformed families demonstrated that a single complete copy of the construct had integrated independently in each case and that it had done so in a transposase-mediated manner. The availability of a mariner transformation system greatly enhances our ability to study and manipulate this important vector species.The incidence of vector-borne diseases is on the rise. As part of a multifaceted effort to control the transmission of diseases, we are developing tools for the molecular genetic manipulation of mosquitoes (1). We intend to use the tools and techniques of modern molecular biology to generate strains of mosquitoes that are incapable of transmitting a specific pathogen. These strains will be used selectively in release programs to reduce disease transmission. One of the key requirements for this effort is a method for introducing genes into mosquitoes. Recently, the Hermes transposable element from the housefly, Musca domestica, was shown to integrate into the germ line of the yellow fever mosquito, Aedes aegypti (2). The identification of Hermes as a viable candidate for mosquito transgenesis resulted from a strategy that first evaluated the ability of the element to mobilize (excise and insert) in the embryonic soma of the mosquito (3), followed by demonstration that it would integrate into the germ line (2). By using this approach, we show that a modified mariner transposable element efficiently and stably integrates into the germ line of Ae. aegypti.Transposition assays based on the mobilization of a marked transposon from a donor to a target plasmid (4) were used to show that the mariner element, Mos1, from Drosophila mauritiana (5), was capable of mobility in embryos of Ae. aegypti. Subsequently, a genetic transformation experiment showed that Mos1 could integrate into the germ line of the mosquito. This experiment exploited the recently demonstrated ability of a wild-type copy of the Drosophila melanogaster cinnabar (cn ϩ ) gene to complement the white-eye phenotype of the kynurenine hydroxylase-white (kh w ) strain of Ae. aegypti (6-8). We report the successful generation of transgenic Ae. aegypti lines that contai...

nanos gene control DNA mediates developmentally regulated transposition in the yellow fever mosquito Aedes aegypti

“…In total, the 1,676 embryos injected resulted in 440 survivors (26%) and an average transformation frequency of 3.6%. These results are similar to those obtained when helper plasmids are used (7,20). Importantly, no secondary integrations were detected in control animals injected with the EGFP test constructs without helper plasmids.…”

“…These strategies benefit from the availability of transgenesis technologies, and stable integration of engineered genes into A. aegypti, using transposable elements (TEs) (7,8) is now routine (9). Population replacement strategies also depend on the development of effector genes to interfere with and prevent pathogen transmission.…”

Bee Venom Phospholipase Inhibits Malaria Parasite Development in Transgenic Mosquitoes

“…The ingested blood meal (BM) is also visible. and 4% for the following combinations: A. stephensi with piggyBac (14), A. aegypti with Hermes (5, 15, 16), and A. aegypti with mariner (5,17).…”

In vivo transposon mutagenesis of the methanogenic archaeon Methanosarcina acetivorans C2A using a modified version of the insect mariner -family transposable element Himar1

“…Thus, in vitro transposition reactions containing only target DNA, transposon DNA, and purified mariner transposase catalyze cut-and-paste transposition in a very efficient manner. This feature has allowed mariner elements to be used for mutagenesis of a wide variety of non-native organisms, including other insects (22), zebrafish (23), protozoa (24), chickens (25), and human cells (in culture) (26). Even more impressive, versions of the element Himar1, originally isolated from the hornfly, Hematobia irritans, that express the transposase gene from appropriate bacterial promoters function in E. coli and Mycobacterium smegmatis (27).…”