Efficient Targeted Integration at leu1-32 and ura4-294 in Schizosaccharomyces pombe

INVESTIGATIONS

Efficient Targeted Integration at leu1-32 and ura4-294 in Schizosaccharomyces pombe

J. B. Keeney and J. D. Boeke
Johns Hopkins University School of Medicine, Department of Molecular Biology and Genetics, Baltimore, Maryland 21205

Homologous integration into the fission yeast Schizosaccharomyces pombe has not been well characterized. In this study, we have examined integration of plasmids carrying the leu1(+) and ura4(+) genes into their chromosomal loci. Genomic DNA blot analysis demonstrated that the majority of transformants have one or more copies of the plasmid vector integrated via homologous recombination with a much smaller fraction of gene conversion to leu1(+) or ura4(+). Non-homologous recombination events were not observed for either gene. We describe the construction of generally useful leu1(+) and ura4(+) plasmids for targeted integration at the leu1-32 and ura4-294 loci of S. pombe.

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[

				M. Sanso, M. Gogol, J. Ayte, C. Seidel, and E. Hidalgo Transcription Factors Pcr1 and Atf1 Have Distinct Roles in Stress- and Sty1-Dependent Gene Regulation Eukaryot. Cell, May 1, 2008; 7(5): 826 - 835. [Abstract] [Full Text] [PDF]

				L. Perez-Hidalgo, A. E. Rozalen, C. Martin-Castellanos, and S. Moreno Slk1 is a meiosis-specific Sid2-related kinase that coordinates meiotic nuclear division with growth of the forespore membrane J. Cell Sci., May 1, 2008; 121(9): 1383 - 1392. [Abstract] [Full Text] [PDF]

				D. M. Clifford, B. A. Wolfe, R. H. Roberts-Galbraith, W. H. McDonald, J. R. Yates III, and K. L. Gould The Clp1/Cdc14 phosphatase contributes to the robustness of cytokinesis by association with anillin-related Mid1 J. Cell Biol., April 3, 2008; 181(1): 79 - 88. [Abstract] [Full Text] [PDF]

				A. Mercier, S. Watt, J. Bahler, and S. Labbe Key Function for the CCAAT-Binding Factor Php4 To Regulate Gene Expression in Response to Iron Deficiency in Fission Yeast Eukaryot. Cell, March 1, 2008; 7(3): 493 - 508. [Abstract] [Full Text] [PDF]

				M. E. Griffith, U. Mayer, A. Capron, Q. A. Ngo, A. Surendrarao, R. McClinton, G. Jurgens, and V. Sundaresan The TORMOZ Gene Encodes a Nucleolar Protein Required for Regulated Division Planes and Embryo Development in Arabidopsis PLANT CELL, July 1, 2007; 19(7): 2246 - 2263. [Abstract] [Full Text] [PDF]

				V. A. Tallada, A. J. Bridge, P. A. Emery, and I. M. Hagan Suppression of the Schizosaccharomyces pombe cut12.1 Cell-Cycle Defect by Mutations in cdc25 and Genes Involved in Transcriptional and Translational Control Genetics, May 1, 2007; 176(1): 73 - 83. [Abstract] [Full Text] [PDF]

				A. Mercier, B. Pelletier, and S. Labbe A Transcription Factor Cascade Involving Fep1 and the CCAAT-Binding Factor Php4 Regulates Gene Expression in Response to Iron Deficiency in the Fission Yeast Schizosaccharomyces pombe Eukaryot. Cell, November 1, 2006; 5(11): 1866 - 1881. [Abstract] [Full Text] [PDF]

				H.-J. Yoon, A. Feoktistova, J.-S. Chen, J. L. Jennings, A. J. Link, and K. L. Gould Role of Hcn1 and Its Phosphorylation in Fission Yeast Anaphase-promoting Complex/Cyclosome Function J. Biol. Chem., October 27, 2006; 281(43): 32284 - 32293. [Abstract] [Full Text] [PDF]

				J. A. Rosenberg, G. C. Tomlin, W. H. McDonald, B. E. Snydsman, E. G. Muller, J. R. Yates III, and K. L. Gould Ppc89 Links Multiple Proteins, Including the Septation Initiation Network, to the Core of the Fission Yeast Spindle-Pole Body Mol. Biol. Cell, September 1, 2006; 17(9): 3793 - 3805. [Abstract] [Full Text] [PDF]

				S. A. Rincon, B. Santos, and P. Perez Fission Yeast Rho5p GTPase Is a Functional Paralogue of Rho1p That Plays a Role in Survival of Spores and Stationary-Phase Cells Eukaryot. Cell, March 1, 2006; 5(3): 435 - 446. [Abstract] [Full Text] [PDF]

				J. Beaudoin and S. Labbe Copper Induces Cytoplasmic Retention of Fission Yeast Transcription Factor Cuf1 Eukaryot. Cell, February 1, 2006; 5(2): 277 - 292. [Abstract] [Full Text] [PDF]

				J. Beaudoin, J. Laliberte, and S. Labbe Functional dissection of Ctr4 and Ctr5 amino-terminal regions reveals motifs with redundant roles in copper transport Microbiology, January 1, 2006; 152(1): 209 - 222. [Abstract] [Full Text] [PDF]

				C. S. Petit, S. Mehta, R. H. Roberts, and K. L. Gould Ace2p contributes to fission yeast septin ring assembly by regulating mid2+ expression J. Cell Sci., December 15, 2005; 118(24): 5731 - 5742. [Abstract] [Full Text] [PDF]

				J. L. Morrell-Falvey, L. Ren, A. Feoktistova, G. D. Haese, and K. L. Gould Cell wall remodeling at the fission yeast cell division site requires the Rho-GEF Rgf3p J. Cell Sci., December 1, 2005; 118(23): 5563 - 5573. [Abstract] [Full Text] [PDF]

				E. B. Gomez, J. M. Espinosa, and S. L. Forsburg Schizosaccharomyces pombe mst2+ Encodes a MYST Family Histone Acetyltransferase That Negatively Regulates Telomere Silencing Mol. Cell. Biol., October 15, 2005; 25(20): 8887 - 8903. [Abstract] [Full Text] [PDF]

				B. Pelletier, A. Trott, K. A. Morano, and S. Labbe Functional Characterization of the Iron-regulatory Transcription Factor Fep1 from Schizosaccharomyces pombe J. Biol. Chem., July 1, 2005; 280(26): 25146 - 25161. [Abstract] [Full Text] [PDF]

				A. Bimbo, J. Liu, and M. K. Balasubramanian Roles of Pdk1p, a Fission Yeast Protein Related to Phosphoinositide-dependent Protein Kinase, in the Regulation of Mitosis and Cytokinesis Mol. Biol. Cell, July 1, 2005; 16(7): 3162 - 3175. [Abstract] [Full Text] [PDF]

				A. P. Vivancos, E. A. Castillo, B. Biteau, C. Nicot, J. Ayte, M. B. Toledano, and E. Hidalgo A cysteine-sulfinic acid in peroxiredoxin regulates H2O2-sensing by the antioxidant Pap1 pathway PNAS, June 21, 2005; 102(25): 8875 - 8880. [Abstract] [Full Text] [PDF]

				A. Bimbo, Y. Jia, S. L. Poh, R. K. M. Karuturi, N. den Elzen, X. Peng, L. Zheng, M. O'Connell, E. T. Liu, M. K. Balasubramanian, et al. Systematic Deletion Analysis of Fission Yeast Protein Kinases Eukaryot. Cell, April 1, 2005; 4(4): 799 - 813. [Abstract] [Full Text] [PDF]

				D. G. Pankratz and S. L. Forsburg Meiotic S-Phase Damage Activates Recombination without Checkpoint Arrest Mol. Biol. Cell, April 1, 2005; 16(4): 1651 - 1660. [Abstract] [Full Text] [PDF]

				V. Tajadura, B. Garcia, I. Garcia, P. Garcia, and Y. Sanchez Schizosaccharomyces pombe Rgf3p is a specific Rho1 GEF that regulates cell wall {beta}-glucan biosynthesis through the GTPase Rho1p J. Cell Sci., December 1, 2004; 117(25): 6163 - 6174. [Abstract] [Full Text] [PDF]

				H. An, J. L. Morrell, J. L. Jennings, A. J. Link, and K. L. Gould Requirements of Fission Yeast Septins for Complex Formation, Localization, and Function Mol. Biol. Cell, December 1, 2004; 15(12): 5551 - 5564. [Abstract] [Full Text] [PDF]

				M. G. Ferreira and J. P. Cooper Two modes of DNA double-strand break repair are reciprocally regulated through the fission yeast cell cycle Genes & Dev., September 15, 2004; 18(18): 2249 - 2254. [Abstract] [Full Text] [PDF]

				L.-L. Du, B. A. Moser, and P. Russell Homo-oligomerization Is the Essential Function of the Tandem BRCT Domains in the Checkpoint Protein Crb2 J. Biol. Chem., September 10, 2004; 279(37): 38409 - 38414. [Abstract] [Full Text] [PDF]

				S. J. Radford, M. L. Boyle, C. J. Sheely, J. Graham, D. P. Haeusser, L. Zimmerman, and J. B. Keeney Increase in Ty1 cDNA Recombination in Yeast sir4 Mutant Strains at High Temperature Genetics, September 1, 2004; 168(1): 89 - 101. [Abstract] [Full Text] [PDF]

				K. Tanaka and P. Russell Cds1 Phosphorylation by Rad3-Rad26 Kinase Is Mediated by Forkhead-associated Domain Interaction with Mrc1 J. Biol. Chem., July 30, 2004; 279(31): 32079 - 32086. [Abstract] [Full Text] [PDF]

				A. Kita, R. Sugiura, H. Shoji, Y. He, L. Deng, Y. Lu, S. O. Sio, K. Takegawa, M. Sakaue, H. Shuntoh, et al. Loss of Apm1, the {micro}1 Subunit of the Clathrin-Associated Adaptor-Protein-1 Complex, Causes Distinct Phenotypes and Synthetic Lethality with Calcineurin Deletion in Fission Yeast Mol. Biol. Cell, June 1, 2004; 15(6): 2920 - 2931. [Abstract] [Full Text] [PDF]

				M.-C. Hou, D. A. Guertin, and D. McCollum Initiation of Cytokinesis Is Controlled through Multiple Modes of Regulation of the Sid2p-Mob1p Kinase Complex Mol. Cell. Biol., April 15, 2004; 24(8): 3262 - 3276. [Abstract] [Full Text] [PDF]

				B. A. Weir and M. P. Yaffe Mmd1p, a Novel, Conserved Protein Essential for Normal Mitochondrial Morphology and Distribution in the Fission Yeast Schizosaccharomyces pombe Mol. Biol. Cell, April 1, 2004; 15(4): 1656 - 1665. [Abstract] [Full Text] [PDF]

				E. Oltra, F. Verde, R. Werner, and G. D'Urso A novel RING-finger-like protein Ini1 is essential for cell cycle progression in fission yeast J. Cell Sci., February 22, 2004; 117(6): 967 - 974. [Abstract] [Full Text] [PDF]