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Stability of Human Centromeric Alphoid DNA Repeat during Propagation in Recombination-Deficient Yeast Strains  

Kim, Kwang-Sup (Department of Biological Science, Dong-A University)
Shin, Young-Sun (Department of Biological Science, Dong-A University)
Lee, Sang-Yeop (Department of Biological Science, Dong-A University)
Ahn, Eun-Kyung (Department of Biological Science, Dong-A University)
Do, Eun-Ju (Department of Biological Science, Dong-A University)
Park, In-Ho (Department of Biological Science, Dong-A University)
Leem, Sun-Hee (Department of Biological Science, Dong-A University)
SunWoo, Yang-Il (Department of Biological Science, Dong-A University)
Publication Information
Korean Journal of Microbiology / v.43, no.4, 2007 , pp. 243-249 More about this Journal
Abstract
The centromere is a highly differentiated structure of the chromosome that fulfills a multitude of essential mitotic and meiotic functions. Alphoid DNA (${\alpha}$-satellite) is the most abundant family of repeated DNA found at the centromere of all human chromosomes, and chromosomes of primates in general. The most important parts in the development of Human Artificial Chromosomes (HACs), are the isolation and maintenance of stability of centromeric region. For isolation of this region, we could use the targeting hook with alphoid DNA repeat and cloned by Transformation-Associated Recombination (TAR) cloning technique in yeast Saccharomyces cerevisiae. The method includes rolling-circle amplification (RCA) of repeats in vitro to 5 kb-length and elongation of the RCA products by homologous recombination in yeast. Four types of $35\;kb{\sim}50\;kb$ of centromeric DNA repeat arrays (2, 4, 5, 6 mer) are used to examine the stability of repeats in homologous recombination mutant strains (rad51, rad52, and rad54). Following the transformation into wild type, rad51 and rad54 mutant strains, there were frequent changes in inserted size. A rad52 mutant strain showed extremely low transformation frequency, but increased stability of centromeric DNA repeat arrays at least 3 times higher than other strains. Based on these results, the incidence of large mutations could be reduced using a rad52 mutant strain in maintenance of centromeric DNA repeat arrays. This genetic method may use more general application in the maintenance of tandem repeats in construction of HAC.
Keywords
alphoid DNA repeats array; homologous recombination; Saccharomyces cerevisiae; TAR cloning;
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1 De Lange, T. 2004. T-loops and the origin of telomeres. Nature Rev. Mol. Cell. Biol. 5, 323-329   DOI   ScienceOn
2 Ebersole, T., Y. Okamoto, V.N. Noskov, N. Kouprina, J.H. Kim, S.H. Leem, J.C. Barrett, H. Masumoto, and V. Larionov. 2005. Rapid generation of long synthetic tandem repeats and its application for analysis in human artificial chromosome formation. Nucleic Acids Res. 33, e130   DOI   ScienceOn
3 Lee, C., R. Wevrick, R.B. Fisher, M.A. Ferguson-Smith, and C.C. Lin. 1997. Human centromeric DNAs. Hum. Genet. 100, 291-304   DOI
4 Noskov, V.N., M. Koriabine, G. Solomone, M. Randolph, J.C. Barrett, S.H. Leem, L. Stubbs, N. Kouprina, and V. Larionov. 2002. Defining the minimal length of sequence homology required for selective gene isolation by TAR cloning. Nucleic Acids Res. 29, e32
5 Sambrook, J., E.F. Fritsch, and T. Maniatis. 1989. Molecular cloning: a laboratory manual, 2nd (ed), Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., USA
6 Sikorski, R. and P. Philip. 1989. A system of shuttle vectors and yeast host strains designed for efficient manipulation of DNA in Saccharomyces cerevisiae. Genetics 122, 19-27   PUBMED
7 Kouprina, N., S.H. Leem, G. Solomon, A. Ly, M. Koriabine, J. Otstot, E. Pak, A. Dutra, S. Zhao, J.C. Barrett, and V. Larionov. 2003. Segments missing from the draft human genome sequence can be isolated by TAR cloning in yeast. EMBO Rep. 4, 257-262   DOI   ScienceOn
8 Razin, S.V., E.S. Ioudinkova, E. Trifonov, and K. Scherrer. 2001. Non-clonability correlates with genome instability: A case of unique DNA region. J. Mol. Biol. 307, 481-486   DOI   ScienceOn
9 Green, E.D., H.C. Reithman, J.E. Dutchik, and M.V. Olson. 1991. Detection and characterization of chimeric yeast artificial chromosome clones. Genomics 11, 658-669   DOI
10 Kouprina, N. and V. Larionov. 1999. Selective Isolation of mammalian Genes by TAR cloning. In A.L. Boyle. (ed.), Current Protocols in Human Genetics, UNIT 5.17, John Wiley & Sons, Inc. New York, NY, USA
11 Schroth, G.P. and P.S. Ho. 1995. Occurrence of potential cruciform and H-DNA forming sequences in genomic DNA. Nucleic Acids Res. 23, 1977-1983   DOI
12 Neil, D.L., A. Villasante, R.B. Fisher, D. Vetrie, B. Cox, and C. Tyler-Smith. 1990. Structural instability of human tandemly repeated DNA sequences cloned in yeast artificial chromosome vectors. Nucleic Acids Res. 18, 1421-1428   DOI
13 Leem, S.H., V.N. Noskov, J.E. Park, S.I. Kim, V. Larionov, and N. Kouprina. 2003. Optimum conditions for selective isolation of genes from complex genomes by transformation-associated recombination cloning. Nucleic Acid Res. 31, e29   DOI   ScienceOn
14 Sherman, F., G.R. Fink, and J.B. Hicks. 1986. Methods in yeast genetics. Cold Spring Harbor Laboratory Press, Cold Spring Harbor, N.Y., USA
15 Noskov, V.N., S.H. Leem, G. Solomone, M. Mullokandov, J.Y. Chae, Y.H. Yoon, Y.S. Shin, N. Kouprina, and V. Larionov. 2003. A novel strategy for analysis of gene homologues and segmental genome duplications. J. Mol. Evol. 56, 702-710   DOI
16 Altschul, S.F., W. Gish, W. Miller, E. Mayers, and D.J. Lipman. 1990. Basic local alignment search tool. J. Mol. Biol. 215, 403-410   DOI   PUBMED
17 Kang, H.K. and D.W. Cox. 1996. Tandem repeats 3' of the IGHA genes in the human immunoglobulin heavy chain gene cluster. Genomics 35, 189-195   DOI   ScienceOn
18 Kim, J.H., Y.S. Shin, Y.H. Yoon, H.J. Jang, E.A. Kim, K.S. Kim, C.N. Chung, I.H. Park, S.H. Leem, and Y. Sunwoo. 2003. Effect of GC content on target hook required for gene isolation by transformation- associated recombination cloning. Kor. J. Microbiol. 39, 128-134
19 Larionov, V., J. Graves, N. Kouprina, and M.A. Resnick. 1994. The role of recombination and RAD52 in mutation of chromosomal DNA transformed into yeast. Nucleic Acid Res. 22, 4234-4241   DOI   ScienceOn
20 Petek, E., P.M. Kroisel, and K. Wagner. 1997. Isolation of site-specific insert probes from chimeric YACs. BioTechniques 23, 72-77   PUBMED
21 Leem, S.H., N. Kouprina, J. Grimwood, J.H. Kim, M. Mullokandov, Y.H. Yoon, J.Y. Chae, J. Morgan, S. Lucas, P. Richardson, C. Detter, T. Glavina, E. Rubin, J.C. Barrett, and V. Larionov. 2004. Closing the gaps on human chromosome 19 revealed genes with a high density of repetitive tandemly arrayed elements. Genome Res. 14, 239-246   DOI   ScienceOn
22 Hagan, C.E. and G.J. Warren. 1982. Lethality of palindromic DNA and its use in selection of recombinant plasmids. Gene 19, 147-151   DOI   ScienceOn