Skip to main content

Thank you for visiting nature.com. You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

  • Research Article
  • Published:

Construction of YAC–based mammalian artificial chromosomes

Nature Biotechnology volume 16, pages 431–439 (1998)Cite this article

Abstract

To construct a mammalian artificial chromosome (MAC), telomere repeats and selectable markers were introduced into a 100 kb yeast artificial chromosome (YAC) containing human centromeric DNA. This YAC, which has a regular repeat structure of alpha-satellite DNA and centromere protein B (CENP-B) boxes, efficiently formed MACs that segregated accurately and bound CENP-B, CENP-C, and CENP-E. The MACs appear to be about 1–5 Mb in size and contain YAC multimers. Structural analyses suggest that the MACs have not acquired host sequences and were formed by a de novo mechanism. The accurate segregation of the MACs suggests they have potential as vectors for introducing genes into mammals.

This is a preview of subscription content, access via your institution

Access options

Buy this article

  • Purchase on Springer Link
  • Instant access to full article PDF
Buy now

Prices may be subject to local taxes which are calculated during checkout

Similar content being viewed by others

References

  1. Murray, A.W. and Szostak, J.W. 1983. Construction of artificial chromosomes in yeast. Nature 305: 189–193.

    Article  CAS  Google Scholar 

  2. Burke, D.T., Carle, G.F., and Olson, M.V. 1987. Cloning of large segments of exogenous DNA into yeast by means of artificial chromosome vectors. Science 236: 806–812.

    Article  CAS  Google Scholar 

  3. Jakobovits, A., Moore, A.L., Green, L.L., Vergara, G.J., Maynard-Currie, C.E., Austin, H.A., and Klapholz, S. 1993. Germ-line transmission and expression of a human-derived yeast artificial chromosome. Nature 362: 255–258.

    Article  CAS  Google Scholar 

  4. Hadlaczky, G., Praznovzky, T., Cserpan, I., Kereso, J., Peterfy, M., Kelemen, I. et al. 1991. Centromere formation in mouse cells cotransformed with human DNA and a dominant marker gene. Proc. Natl. Acad. Sci. USA 88: 8106–8110.

    Article  CAS  Google Scholar 

  5. Sun, T.-Q., Fenstermacher, D.A., and Vos, J.M.H. 1994. Human artificial episomal chromosomes for cloning large DNA fragments in human cells. Nat. Gen. 8: 33–41.

    Article  CAS  Google Scholar 

  6. Farr, C.J., Bayne, R.A.L., Kipling, D., Mills, W., Critcher, R., and Cooke, H.J. 1995. Generation of a human X-derived minichromosome using telomere-associated chromosome fragmentation. EMBO J. 14: 5444–5454.

    Article  CAS  Google Scholar 

  7. Heller, R., Brown, K.E., Burgtorf, C. and Brown, W.R.A. 1996. Mini-chromosome derived from the human Y chromosome by telomere directed chromosome breakage. Proc. Natl. Acad. Sci. USA 93: 7125–7130.

    Article  CAS  Google Scholar 

  8. Brown, W.R.A. 1989. Molecular cloning of human telmomeres in yeast. Nature 338: 774–776.

    Article  CAS  Google Scholar 

  9. Cross, S.H., Allshire, R.C., McKay, S.J., McGill, N.I., and Cooke, H.J. 1989. Cloning of human telomeres by complementation in yeast. Nature 338: 771–774.

    Article  CAS  Google Scholar 

  10. Farr, C.J., Fantes, J., Goodfellow, P. and Cooke, H.J. 1991. Functional reintro-duction of human telomeres into mammalian cells. Proc. Natl. Acad. Sci. USA 88: 7006–7010.

    Article  CAS  Google Scholar 

  11. Barnett, M.A., Buckle, V.J., Evans, E.P., Porter, A.C.G., Rout, D., Smith, A.G., and Brown, W.R.A. 1993. Telomere directed fragmentation of mammalian chromosomes. Nucl. Acids Res. 21: 27–36.

    Article  CAS  Google Scholar 

  12. Willard, H.F, and Waye, J.S. 1987. Hierarchical order in chromosome-specific human alpha satellite DNA. Trends Genet. 3: 192–198.

    Article  CAS  Google Scholar 

  13. Choo, K.H., Vissel, B., Nagy, A., Earle, E., and Kalitsis, P. 1991. A survey of the genomic distribution of alpha satellite DNA on all the human chromosomes, and derivation of a new consensus sequence. Nucleic Acids Res. 19:1179–1182.

    Article  CAS  Google Scholar 

  14. Masumoto, H., Sugimoto, K., and Okazaki, T. 1989. Alphoid satellite DNA is tightly associated with centromere antigens in human chromosome throughout the cell cell cycle. Exp. Cell Res. 181: 181–196.

    Article  CAS  Google Scholar 

  15. Palmer, D.K., O'Day, K., Trong, H.L., Charbonneau, H., and Margolis, R.L. 1991. Purification of the centromere-specific protein CENP-A and demonstration that it is a distinctive histone. Proc. Natl. Acad. Sci. USA 88: 3734–3738.

    Article  CAS  Google Scholar 

  16. Masumoto, H., Masukata, H., Muro, Y., Nozaki, N., and Okazaki, T. 1989. A human centromere antigen (CENP-B) interacts with a short specific sequence in alphoid DNA, a human centromeric satellite. J. Cell Biol. 109: 1963–1973.

    Article  CAS  Google Scholar 

  17. Kitagawa, K., Masumoto, H., Ikeda, M. and Okazaki, T. 1995. Analysis of protein-DNA and protein-protein interactions of centromere protein B (CENP-B) and properties of the DNA-CENP-B complex in the cell cycle. Mol. Cell. Biol. 15: 1602–1612.

    Article  CAS  Google Scholar 

  18. Saitoh, H., Tomkiel, J., Cooke, C.A., Ratire, H., Maurer, M., Rothfield, N.F., and Earnshaw, W.C. 1992. CENP-C, an autoantigen in scleroderma, is a component of the human inner kinetochore plate. Cell 70: 115–125.

    Article  CAS  Google Scholar 

  19. Yen, T.J., Compton, D.A., Wise, D., Zinkowski, R.P., Brinkley, B.R., Earnshaw, W.C., and Cleveland, D.W. 1991. CENP-E, a novel human centromere-associated protein required for progression from metaphase to anaphase. EMBO J. 10: 1245–1254.

    Article  CAS  Google Scholar 

  20. Harrington, J.J., Van Bokkelen, G., Mays, R.W., Gustashaw, K., and Willard, H.F. 1997. Formation of de novo centromeres and construction of first-generation human artificial microchromosomes. Nat. Genet. 4: 345–355.

    Article  Google Scholar 

  21. Neil, D.L., Villasante, A., Fisher, R.B., Vetrie, D., Cox, B., and Tyler-Smith, C. 1990. Structural instability of human tandemly repeated DNA sequences cloned in yeast artificial chromosome vectors. Nucleic Acids Res. 18: 1421–1428.

    Article  CAS  Google Scholar 

  22. Kouprina, N., Eldarov, M., Moyzis, R., Resnick, M., and Larionov, V. 1994. A model system to assess the integrity of mammalian YACs during transformation and propagation in yeast. Genomics 21: 7–17.

    Article  CAS  Google Scholar 

  23. Ikeno, M., Masumoto, H., and Okazaki, T. 1994. Distribution of CENP-B boxes reflected in CREST centromere antigenic sites on long-range a satellite DNA arrays of human chromosome 21. Hum. Mol. Genet. 3: 1245–1257.

    Article  CAS  Google Scholar 

  24. Pavan, W.J., Hieter, P. and Reeves, R.H. 1990. Generation of deletion derivatives by targeted transformation of human-derived yeast artificial chromosomes. Proc. Natl. Acad. Sci. USA 87:1300–1304.

    Article  CAS  Google Scholar 

  25. Adzuma, K., Ogawa, T., and Ogawa, H. 1984. Primary structure of the RAD52 gene in Saccharomyces cerevisiae. Mol. Cell. Biol. 4: 2735–2744.

    Article  CAS  Google Scholar 

  26. Taylor, S.S., Larin, Z., and Tyler-Smith, C. 1996. Analysis of extrachromosomal structures containing human centromeric alphoid satellite DNA sequences in mouse cells. Chromosoma 105: 70–81.

    Article  CAS  Google Scholar 

  27. Earnshaw, W.C., Ratrie, H., and Stetten, G. 1989. Visualization of centromere proteins CENP-B and CENP-C on a stable dicentric chromosome in cytological spreads. Chromosoma 98: 1–12.

    Article  CAS  Google Scholar 

  28. Sullivan, B.A. and Schwartz, S. 1995. Identification of centromeric antigens in dicentric Robertsonian translocations: CENP-C and CENP-E are necessary components of functional centromeres. Hum, Mol. Genet. 4: 2189–2197.

    Article  CAS  Google Scholar 

  29. Krysan, P.J., Smith, J.G., and Calos, M.R. 1993. Autonomous replication in human cells of multimers of specific human and bacterial DNA sequences. Mol. Cell. Biol. 13: 2688–2696.

    Article  CAS  Google Scholar 

  30. Featherstone, T. and Huxley, C. 1993. Extrachromosomal maintenance and amplification of yeast artificial chromosome DNA in mouse cells. Genomics 17: 267–278.

    Article  CAS  Google Scholar 

  31. Muro, Y., Masumoto, H., Yoda, K., Nozaki, N., Ohashi, M. and Okazaki, T. 1992. Centromere protein B assembles human centromeric a-satellite DNA at the 17 bp sequence, CENP-B box. J. Cell Biol. 116: 585–596.

    Article  CAS  Google Scholar 

  32. Masumoto, H., Yoda, K., Ikeno, M., Kitagawa, K., Muro, Y., and Okazaki, T. 1993. Properties of CENP-B and its target sequence in a satellite DNA, pp. 31–43 in Chromosome segregation and aneuploidy. Vig, B.K. (ed.). NATO ASI Series, Springer Verlag, Germany.

    Chapter  Google Scholar 

  33. Haff, T., Mater, A.G., Wienberg, J., and Ward, D.C. 1995. Presence and abundance of CENP-B box sequences in great ape subsets of primate-specific alpha-satellite DNA. J. Mol. Evol. 41: 487–491.

    Article  Google Scholar 

  34. Broccoli, D., Miller, O.J., and Miller, D.A. 1990. Relationship of mouse minor satellite DNA to centromere activity. Cytogenet. Cell Genet. 54: 182–186.

    Article  CAS  Google Scholar 

  35. du Sart, D., Cancilla, M.R., Earle, E., Mao, J., Saffery, R., Tainton, K.M. et al. 1997. A functional neo-centromere formed through activation of a latent human centromere and consisting of non-alpha-satellite DNA. Nat. Genet. 16: 144–153.

    Article  CAS  Google Scholar 

  36. Steiner, N.C. and Clarke, L. 1994. A novel epigenetic effect after centromere function in fission yeast. Cell 79: 865–874.

    Article  CAS  Google Scholar 

  37. Raziuddin, A., Sarkar, F.H., Dutkowski, R., Shulman, L., Ruddle, F.H., and Gupta, S.L. 1984. Receptors for human α and β interferon but not for γ interferon are specified by human chromosome 21. Proc. Natl. Acad. Sci. USA 84: 5504–5508.

    Article  Google Scholar 

  38. Kalitsis, P., Earle, E., Vissel, B., Shaffer, L.G., and Choo, K.H.A. 1993. A chromosome 13-specific human satellite I DNA subfamily with minor presence on chromosome 21: further studies on robertsonian translocations. Genomics 16: 104–112.

    Article  CAS  Google Scholar 

  39. Vissel, B., Nagy, A. and Choo, K.H.A. 1992. A satellite III sequence shared by human chromosome 13, 14, and 21 that is contiguous with alpha satellite DNA.Cytogenet. Cell Genet. 61: 81–86.

    Article  CAS  Google Scholar 

  40. Ledbetter, S.A., Nelson, D.L., Warren, S.T. and Ledbetter, D.H. 1990. Rapid isolation of DNA probes within specific chromosome region by interspersed repetitive sequence polymerase chain reaction. Genomics 6: 475–481.

    Article  CAS  Google Scholar 

  41. Smith, D.R., Smyth, A.P., Strauss, W.M., and Moir, D.T. 1993. Incorporation of copy-number control elements into yeast artificial chromosomes by targeted homologous recombination. Mamm. Genome 4: 141–147.

    Article  CAS  Google Scholar 

  42. Cross, S.H. Isolation and characterisation of human telomeres. (Ph.D. Thesis, University of Edinburgh, Scotland, 1989).

  43. Shero, J.H., McCormick, M.K., Antonarakis, S., and Hieter, P. 1991. Yeast artificial chromosome vectors for efficient clone manipulation and mapping. Genomics 10: 505–508.

    Article  CAS  Google Scholar 

  44. Izumi, M., Yazawa, H., Kamakura, T., Yamaguchi, I., Endo, T., and Hanaoka, F. 1991. S-resistance gene (bsr): a novel selectable marker for mammalian cells. Exp. Cell Res. 197: 229–233.

    Article  CAS  Google Scholar 

  45. Schedl, A., Grimes, B., and Montoliu, L. 1995. YAC-transfer by microinjection, pp. 281–306 in Methods in molecular biology. Markie, D. (ed.). Humana Press, Totowa, NJ.

    Google Scholar 

  46. Muramatsu, M., Yamamoto, O., Kishimoto, T.M.N., Kato, H., and Kominami, R. 1986. Structure and regulation of mammalian ribosomal RNA gene. Adv. Biophys. 21:217–227.

    Article  CAS  Google Scholar 

  47. Lansdorp, P.M., Verwoerd, N.P., van de Rijke, F.M., Dragowska, V., Little, M.T., Dirks, R.W. et al. 1996. Heterogeneity in telomere length of human chromosomes. Hum. Mol. Genet. 5: 685–691.

    Article  CAS  Google Scholar 

  48. Trowell, H.E., Nagy, A., Vissel, B., and Choo, K.H.A. 1993. Long-range analyses of the centromeric regions of human chromosome 13,14 and 21: identification of a narrow domain containing two key centromeric DNA elements. Hum. Mol. Genet. 2: 1639–1649.

    Article  CAS  Google Scholar 

Download references

Author information

Author notes

  1. Masashi Ikeno & Tuneko Okazaki

    Present address: Institute for Comprehensive Medical Science, Fujita Health University, Toyoake, Aichi, 470-1101, Japan

  2. Hiroshi Masumoto: Corresponding author e-mail: g44478a@nucc.cc.nagoya-u.ac.jp

Authors and Affiliations

  1. Department of Molecular Biology, School of Science, Nagoya University, Chikusa-ku, Nagoya, 464-8602, Japan

    Masashi Ikeno, Tuneko Okazaki, Megumi Nakano, Kaori Saitoh, Harumi Hoshino & Hiroshi Masumoto

  2. MRC Human Genetics Unit, Western General Hospital, Edinburgh, EH4 2XU, UK

    Brenda Grimes, Niolette I. McGill & Howard Cooke

Authors
  1. Masashi Ikeno
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Brenda Grimes
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Tuneko Okazaki
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Megumi Nakano
    View author publications

    You can also search for this author in PubMed Google Scholar

  5. Kaori Saitoh
    View author publications

    You can also search for this author in PubMed Google Scholar

  6. Harumi Hoshino
    View author publications

    You can also search for this author in PubMed Google Scholar

  7. Niolette I. McGill
    View author publications

    You can also search for this author in PubMed Google Scholar

  8. Howard Cooke
    View author publications

    You can also search for this author in PubMed Google Scholar

  9. Hiroshi Masumoto
    View author publications

    You can also search for this author in PubMed Google Scholar

Rights and permissions

Reprints and permissions

About this article

Cite this article

Ikeno, M., Grimes, B., Okazaki, T. et al. Construction of YAC–based mammalian artificial chromosomes. Nat Biotechnol 16, 431–439 (1998). https://doi.org/10.1038/nbt0598-431

Download citation

  • Received:

  • Accepted:

  • Issue Date:

  • DOI: https://doi.org/10.1038/nbt0598-431

This article is cited by

Search

Advanced search

Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing