Abstract
The ability to place a series of gene constructs at a specific site in the genome opens new possibilities for the experimental examination of gene expression and chromosomal position effects. We report that the FLP- FRT site-specific recombination system of the yeast 2mu plasmid can be used to integrate DNA at a chromosomal FRT target site in Drosophila. The technique we used was to first integrate an FRT- flanked gene by standard P element-mediated transformation. FLP was then used to excise the FRT- flanked donor DNA and screen for FLP-mediated re-integration at an FRT target at a different chromosome location. Such events were recovered from up to 5% of the crosses used to screen for mobilization and are easily detectable by altered linkage of a white reporter gene or by the generation of a white + gene upon integration.
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- Ahmad K., Golic K. G. Somatic reversion of chromosomal position effects in Drosophila melanogaster. Genetics. 1996 Oct;144(2):657–670. doi: 10.1093/genetics/144.2.657. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Albert H., Dale E. C., Lee E., Ow D. W. Site-specific integration of DNA into wild-type and mutant lox sites placed in the plant genome. Plant J. 1995 Apr;7(4):649–659. doi: 10.1046/j.1365-313x.1995.7040649.x. [DOI] [PubMed] [Google Scholar]
- Banga S. S., Boyd J. B. Oligonucleotide-directed site-specific mutagenesis in Drosophila melanogaster. Proc Natl Acad Sci U S A. 1992 Mar 1;89(5):1735–1739. doi: 10.1073/pnas.89.5.1735. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Capecchi M. R. The new mouse genetics: altering the genome by gene targeting. Trends Genet. 1989 Mar;5(3):70–76. doi: 10.1016/0168-9525(89)90029-2. [DOI] [PubMed] [Google Scholar]
- Chalfie M., Tu Y., Euskirchen G., Ward W. W., Prasher D. C. Green fluorescent protein as a marker for gene expression. Science. 1994 Feb 11;263(5148):802–805. doi: 10.1126/science.8303295. [DOI] [PubMed] [Google Scholar]
- Chou T. B., Perrimon N. Use of a yeast site-specific recombinase to produce female germline chimeras in Drosophila. Genetics. 1992 Jul;131(3):643–653. doi: 10.1093/genetics/131.3.643. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Falco S. C., Li Y., Broach J. R., Botstein D. Genetic properties of chromosomally integrated 2 mu plasmid DNA in yeast. Cell. 1982 Jun;29(2):573–584. doi: 10.1016/0092-8674(82)90173-8. [DOI] [PubMed] [Google Scholar]
- Golic K. G., Golic M. M. Engineering the Drosophila genome: chromosome rearrangements by design. Genetics. 1996 Dec;144(4):1693–1711. doi: 10.1093/genetics/144.4.1693. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Golic K. G., Lindquist S. The FLP recombinase of yeast catalyzes site-specific recombination in the Drosophila genome. Cell. 1989 Nov 3;59(3):499–509. doi: 10.1016/0092-8674(89)90033-0. [DOI] [PubMed] [Google Scholar]
- Golic M. M., Golic K. G. A quantitative measure of the mitotic pairing of alleles in Drosophila melanogaster and the influence of structural heterozygosity. Genetics. 1996 May;143(1):385–400. doi: 10.1093/genetics/143.1.385. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Huang L. C., Wood E. A., Cox M. M. A bacterial model system for chromosomal targeting. Nucleic Acids Res. 1991 Feb 11;19(3):443–448. doi: 10.1093/nar/19.3.443. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Keeler K. J., Dray T., Penney J. E., Gloor G. B. Gene targeting of a plasmid-borne sequence to a double-strand DNA break in Drosophila melanogaster. Mol Cell Biol. 1996 Feb;16(2):522–528. doi: 10.1128/mcb.16.2.522. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Klemenz R., Weber U., Gehring W. J. The white gene as a marker in a new P-element vector for gene transfer in Drosophila. Nucleic Acids Res. 1987 May 26;15(10):3947–3959. doi: 10.1093/nar/15.10.3947. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Michiels F., Gasch A., Kaltschmidt B., Renkawitz-Pohl R. A 14 bp promoter element directs the testis specificity of the Drosophila beta 2 tubulin gene. EMBO J. 1989 May;8(5):1559–1565. doi: 10.1002/j.1460-2075.1989.tb03540.x. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Mismer D., Rubin G. M. Analysis of the promoter of the ninaE opsin gene in Drosophila melanogaster. Genetics. 1987 Aug;116(4):565–578. doi: 10.1093/genetics/116.4.565. [DOI] [PMC free article] [PubMed] [Google Scholar]
- O'Gorman S., Fox D. T., Wahl G. M. Recombinase-mediated gene activation and site-specific integration in mammalian cells. Science. 1991 Mar 15;251(4999):1351–1355. doi: 10.1126/science.1900642. [DOI] [PubMed] [Google Scholar]
- Orr-Weaver T. L., Szostak J. W., Rothstein R. J. Genetic applications of yeast transformation with linear and gapped plasmids. Methods Enzymol. 1983;101:228–245. doi: 10.1016/0076-6879(83)01017-4. [DOI] [PubMed] [Google Scholar]
- Petersen R. B., Lindquist S. Regulation of HSP70 synthesis by messenger RNA degradation. Cell Regul. 1989 Nov;1(1):135–149. doi: 10.1091/mbc.1.1.135. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Robertson H. M., Preston C. R., Phillis R. W., Johnson-Schlitz D. M., Benz W. K., Engels W. R. A stable genomic source of P element transposase in Drosophila melanogaster. Genetics. 1988 Mar;118(3):461–470. doi: 10.1093/genetics/118.3.461. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Rubin G. M., Spradling A. C. Genetic transformation of Drosophila with transposable element vectors. Science. 1982 Oct 22;218(4570):348–353. doi: 10.1126/science.6289436. [DOI] [PubMed] [Google Scholar]
- Sadowski P. D. The Flp recombinase of the 2-microns plasmid of Saccharomyces cerevisiae. Prog Nucleic Acid Res Mol Biol. 1995;51:53–91. [PubMed] [Google Scholar]
- Sauer B., Henderson N. Targeted insertion of exogenous DNA into the eukaryotic genome by the Cre recombinase. New Biol. 1990 May;2(5):441–449. [PubMed] [Google Scholar]
- Sauer B. Identification of cryptic lox sites in the yeast genome by selection for Cre-mediated chromosome translocations that confer multiple drug resistance. J Mol Biol. 1992 Feb 20;223(4):911–928. doi: 10.1016/0022-2836(92)90252-f. [DOI] [PubMed] [Google Scholar]
- Schlake T., Bode J. Use of mutated FLP recognition target (FRT) sites for the exchange of expression cassettes at defined chromosomal loci. Biochemistry. 1994 Nov 1;33(43):12746–12751. doi: 10.1021/bi00209a003. [DOI] [PubMed] [Google Scholar]
- Seifert H. S., Chen E. Y., So M., Heffron F. Shuttle mutagenesis: a method of transposon mutagenesis for Saccharomyces cerevisiae. Proc Natl Acad Sci U S A. 1986 Feb;83(3):735–739. doi: 10.1073/pnas.83.3.735. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Siegal M. L., Hartl D. L. Transgene Coplacement and high efficiency site-specific recombination with the Cre/loxP system in Drosophila. Genetics. 1996 Oct;144(2):715–726. doi: 10.1093/genetics/144.2.715. [DOI] [PMC free article] [PubMed] [Google Scholar]
- Sun Y. H., Tsai C. J., Green M. M., Chao J. L., Yu C. T., Jaw T. J., Yeh J. Y., Bolshakov V. N. White as a reporter gene to detect transcriptional silencers specifying position-specific gene expression during Drosophila melanogaster eye development. Genetics. 1995 Nov;141(3):1075–1086. doi: 10.1093/genetics/141.3.1075. [DOI] [PMC free article] [PubMed] [Google Scholar]