A transposon based on the transposable element Minos from Drosophila hydei was introduced into the genome of Drosophila melanogaster using transformation mediated by the Minos transposase. The transposon carries a wild-type version of the white gene (w) of Orosophila inserted into the second exon of Minos. Transformation was obtained by injecting the transposon into preblastoderm embryos that were expressing transposase either from a Hsp7O-Minos fusion inserted into the genome via P-element-mediated transformation or from a coinjected plasmid carrying the Hsp7O-Minos fusion. Between 1% and 6% of the fertile injected individuals gave transformed progeny. Four of the insertions were cloned and the DNA sequences flanking the transposon ends were determined. The "empty" sites corresponding to three ofthe insertions were amplified from the recipient strain by PCR, cloned, and sequenced. In all cases, the transposon has inserted into a TA dinucleotide and has created the characteristic TA target site duplication. In the absence of transposase, the insertions were stable in the soma and the germ line. However, in the presence of the Hsp7O-Minos gene the Minos-w transposon excises, resulting in mosaic eyes and germ-line reversion to the white phenotype. Minos could be utilized as an alternative to existing systems for transposon tagging and enhancer trapping in Drosophila; it might also be of use as a germ-line transformation vector for non-Drosophila insects.Transposable elements have been used as vectors for stable germ-line transformation in Drosophila melanogaster. The mobile element P, present in some D. melanogaster populations, was the first element shown to insert into germ-line chromosomes in embryos (1). Typically, P-element-mediated transformation is achieved by coinjecting into preblastoderm embryos a mixture of two plasmids: one expressing transposase but unable to transpose and one carrying a gene (or genes) of interest flanked by the inverted terminal repeats of the element. Transformants are detected among the progeny of the injected individuals through the expression of dominant phenotypes. Germ-line transformation has revolutionized Drosophila research through the introduction of powerful methodologies such as analysis of in vitro mutagenized genes, gene cloning by transposon tagging, and enhancer trapping.Two other elements unrelated to P, hobo and mariner, have been shown to be able to transpose in the genome of D.melanogaster. hobo is found in some but not all D. melanogaster populations and causes hybrid dysgenesis (2, 3). Defective hobo elements containing foreign DNA can transpose into germline chromosomes from plasmids if coinjected into preblastoderm embryos along with a full-length element that can provide transposase (4). mariner, an element found in Drosophila mauritiana but not in D. melanogaster, has been introduced into D. melanogaster and shown to induce in trans mobilization of a nonautonomous mariner element (5, 6). Efforts to transfer the Drosophila germ-line transformation me...

Elements related to the Tcl-like Minos mobile element have been cloned from Drosophila hydei and sequenced. Southern blot and sequence analyses show that (01 the elements are actively transposing in the Drosophila hydei germ line, (it) they are characterized by a striking degree of sequence and size homogeneity, and (ii) like Tcl, they insert at a TA dinucleotide that is probably duplicated during the process. The nucleotide sequences of two elements, Minos-2 and Minos-3, differ at only one position from each other and contain two nonoverlapping open reading frames that are separated by a putative 60-nucleotide intron. The amino-terminal part of the Minos putative transposase shows sequence similarity to the paired DNA-binding domain. Forced transcription of a modified Minos element that was introduced into the Drosophila melanogaster germ line by P element-mediated transformation resulted in the production of accurately spliced polyadenylylated RNA molecules. It is proposed that Minos-2 and/or Minos-3 may encode an active transposase containing an amino-terminal DNA-binding domain that is distantly related to the paired DNA-binding domain.Mobile elements belonging to the Tcl-like family have been identified so far in nematodes, insects, and fish (1-7). They are all characterized by a relatively small length (1.6-1.8 kb), the presence of inverted terminal repeats of various sizes and sequences, and significant sequence similarities in the region between the repeats, which corresponds to the gene encoding transposase.A common characteristic of elements capable of transposing autonomously is the presence of a gene encoding transposase, a protein directly involved in the transposition. With the exception of Tcl, TCb1, and Bari-i, the elements of the family that have been sequenced so far do not encode active transposases, having accumulated nonsense and frameshift mutations in their putative transposase genes. Tcl, a 1611-bp element with 54-bp perfect inverted terminal repeats, contains a gene, TcJA, encoding a transposase that binds specifically at the inverted repeats at the ends of the element and induces transposition of endogenous Tcl elements when overexpressed in Caenorhabditis elegans (8). The Tcl element exhibits a high degree of size and sequence homogeneity, in contrast to other eukaryotic transposons (9-11) that are heterogeneous in size. Another characteristic of Tcl is that it always inserts into a TA sequence, possibly creating a duplication of the dinucleotide in the process (12, 13).Minos has been identified as a dispersed repetitive sequence inserted within the transcribed spacer in one of the repeats of the rDNA locus of Drosophila hydei (6). The element is characterized by 255-bp perfect inverted terminal repeats and the presence of two long nonoverlapping open reading frames (ORFs) on the same strand; the longest of the ORFs (ORF2) shows -=30% sequence identity with TcJA but does not begin with an ATG codon (6). It appears, therefore, that the cloned element represents a defective membe...