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. 1993 Feb 25;21(4):969–975. doi: 10.1093/nar/21.4.969

Activity of yeast FLP recombinase in maize and rice protoplasts.

L A Lyznik 1, J C Mitchell 1, L Hirayama 1, T K Hodges 1
PMCID: PMC309231  PMID: 8451196

Abstract

We have demonstrated that a yeast FLP/FRT site-specific recombination system functions in maize and rice protoplasts. FLP recombinase activity was monitored by reactivation of beta-glucuronidase (GUS) expression from vectors containing the gusA gene inactivated by insertion of two FRTs (FLP recombination targets) and a 1.31 kb DNA fragment. The stimulation of GUS activity in protoplasts cotransformed with vectors containing FRT inactivated gusA gene and a chimeric FLP gene depended on both the expression of the FLP recombinase and the presence and structure of the FRT sites. The FLP enzyme could mediate inter- and intramolecular recombination in plant protoplasts. These results provide evidence that a yeast recombination system can function efficiently in plant cells, and that its performance can be manipulated by structural modification of the FRT sites.

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Selected References

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  1. Andrews B. J., Proteau G. A., Beatty L. G., Sadowski P. D. The FLP recombinase of the 2 micron circle DNA of yeast: interaction with its target sequences. Cell. 1985 Apr;40(4):795–803. doi: 10.1016/0092-8674(85)90339-3. [DOI] [PubMed] [Google Scholar]
  2. Bayley C. C., Morgan M., Dale E. C., Ow D. W. Exchange of gene activity in transgenic plants catalyzed by the Cre-lox site-specific recombination system. Plant Mol Biol. 1992 Jan;18(2):353–361. doi: 10.1007/BF00034962. [DOI] [PubMed] [Google Scholar]
  3. Bradford M. M. A rapid and sensitive method for the quantitation of microgram quantities of protein utilizing the principle of protein-dye binding. Anal Biochem. 1976 May 7;72:248–254. doi: 10.1006/abio.1976.9999. [DOI] [PubMed] [Google Scholar]
  4. Cox M. M. The FLP protein of the yeast 2-microns plasmid: expression of a eukaryotic genetic recombination system in Escherichia coli. Proc Natl Acad Sci U S A. 1983 Jul;80(14):4223–4227. doi: 10.1073/pnas.80.14.4223. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Dale E. C., Ow D. W. Gene transfer with subsequent removal of the selection gene from the host genome. Proc Natl Acad Sci U S A. 1991 Dec 1;88(23):10558–10562. doi: 10.1073/pnas.88.23.10558. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Dennis E. S., Gerlach W. L., Pryor A. J., Bennetzen J. L., Inglis A., Llewellyn D., Sachs M. M., Ferl R. J., Peacock W. J. Molecular analysis of the alcohol dehydrogenase (Adh1) gene of maize. Nucleic Acids Res. 1984 May 11;12(9):3983–4000. doi: 10.1093/nar/12.9.3983. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Frearson E. M., Power J. B., Cocking E. C. The isolation, culture and regeneration of Petunia leaf protoplasts. Dev Biol. 1973 Jul;33(1):130–137. doi: 10.1016/0012-1606(73)90169-3. [DOI] [PubMed] [Google Scholar]
  8. 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]
  9. Halfter U., Morris P. C., Willmitzer L. Gene targeting in Arabidopsis thaliana. Mol Gen Genet. 1992 Jan;231(2):186–193. doi: 10.1007/BF00279790. [DOI] [PubMed] [Google Scholar]
  10. Jayaram M. Two-micrometer circle site-specific recombination: the minimal substrate and the possible role of flanking sequences. Proc Natl Acad Sci U S A. 1985 Sep;82(17):5875–5879. doi: 10.1073/pnas.82.17.5875. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Jefferson R. A., Burgess S. M., Hirsh D. beta-Glucuronidase from Escherichia coli as a gene-fusion marker. Proc Natl Acad Sci U S A. 1986 Nov;83(22):8447–8451. doi: 10.1073/pnas.83.22.8447. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Joyner A. L. Gene targeting and gene trap screens using embryonic stem cells: new approaches to mammalian development. Bioessays. 1991 Dec;13(12):649–656. doi: 10.1002/bies.950131206. [DOI] [PubMed] [Google Scholar]
  13. Lee K. Y., Lund P., Lowe K., Dunsmuir P. Homologous recombination in plant cells after Agrobacterium-mediated transformation. Plant Cell. 1990 May;2(5):415–425. doi: 10.1105/tpc.2.5.415. [DOI] [PMC free article] [PubMed] [Google Scholar]
  14. Lyznik L. A., McGee J. D., Tung P. Y., Bennetzen J. L., Hodges T. K. Homologous recombination between plasmid DNA molecules in maize protoplasts. Mol Gen Genet. 1991 Nov;230(1-2):209–218. doi: 10.1007/BF00290670. [DOI] [PubMed] [Google Scholar]
  15. Lyznik L. A., Ryan R. D., Ritchie S. W., Hodges T. K. Stable co-transformation of maize protoplasts with gusA and neo genes. Plant Mol Biol. 1989 Aug;13(2):151–161. doi: 10.1007/BF00016134. [DOI] [PubMed] [Google Scholar]
  16. 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]
  17. Odell J., Caimi P., Sauer B., Russell S. Site-directed recombination in the genome of transgenic tobacco. Mol Gen Genet. 1990 Sep;223(3):369–378. doi: 10.1007/BF00264442. [DOI] [PubMed] [Google Scholar]
  18. Onouchi H., Yokoi K., Machida C., Matsuzaki H., Oshima Y., Matsuoka K., Nakamura K., Machida Y. Operation of an efficient site-specific recombination system of Zygosaccharomyces rouxii in tobacco cells. Nucleic Acids Res. 1991 Dec 11;19(23):6373–6378. doi: 10.1093/nar/19.23.6373. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Ponce M. R., Micol J. L. PCR amplification of long DNA fragments. Nucleic Acids Res. 1992 Feb 11;20(3):623–623. doi: 10.1093/nar/20.3.623. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Russell S. H., Hoopes J. L., Odell J. T. Directed excision of a transgene from the plant genome. Mol Gen Genet. 1992 Jul;234(1):49–59. doi: 10.1007/BF00272344. [DOI] [PubMed] [Google Scholar]
  21. Senecoff J. F., Bruckner R. C., Cox M. M. The FLP recombinase of the yeast 2-micron plasmid: characterization of its recombination site. Proc Natl Acad Sci U S A. 1985 Nov;82(21):7270–7274. doi: 10.1073/pnas.82.21.7270. [DOI] [PMC free article] [PubMed] [Google Scholar]
  22. Senecoff J. F., Cox M. M. Directionality in FLP protein-promoted site-specific recombination is mediated by DNA-DNA pairing. J Biol Chem. 1986 Jun 5;261(16):7380–7386. [PubMed] [Google Scholar]
  23. Senecoff J. F., Rossmeissl P. J., Cox M. M. DNA recognition by the FLP recombinase of the yeast 2 mu plasmid. A mutational analysis of the FLP binding site. J Mol Biol. 1988 May 20;201(2):405–421. doi: 10.1016/0022-2836(88)90147-7. [DOI] [PubMed] [Google Scholar]

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