Assay of centromere function using a human artificial chromosome

Hiroshi Masumoto, Masashi Ikeno, Megumi Nakano, Tuneko Okazaki, Brenda Grimes, Howard Cooke, Nobutaka Suzuki

Research output: Contribution to journalArticle

59 Scopus citations

Abstract

In order to define a functional human centromere sequence, an artificial chromosome was constructed as a reproducible DNA molecule. Mammalian telomere repeats and a selectable marker were introduced into yeast artificial chromosomes (YACs) containing alphoid DNA from the centromere region of human chromosome 21 in a recombination-deficient yeast host. When these modified YACs were introduced into cultured human cells, a YAC with the alphoid DNA from the α21-I locus, containing CENP-B boxes at a high frequency and a regular repeat array, efficiently formed minichromosomes that were maintained stably in the absence of selection and bound CENP-A, CENP-B, CENP-C and CENP-E. The minichromosomes, 1-5 Mb in size and composed of multimers of the introduced YAC DNA, aligned at metaphase plates and segregated to opposite poles correctly in anaphase. Extensive cytological analyses strongly suggested that the minichromosomes had not acquired host sequences and were formed in all cases by a de novo mechanism. In contrast, minichromosomes were never produced with a modified YAC containing alphoid DNA from the α21-II locus, which contains no CENP-B boxes and has a less regular sequence arrangement. We conclude that α21-I alphoid DNA can induce de novo assembly of active centromere/kinetochore structures on minichromosomes.

Original languageEnglish (US)
Pages (from-to)406-416
Number of pages11
JournalChromosoma
Volume107
Issue number6-7
DOIs
StatePublished - Jan 1 1998
Externally publishedYes

ASJC Scopus subject areas

  • Genetics
  • Genetics(clinical)

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  • Cite this

    Masumoto, H., Ikeno, M., Nakano, M., Okazaki, T., Grimes, B., Cooke, H., & Suzuki, N. (1998). Assay of centromere function using a human artificial chromosome. Chromosoma, 107(6-7), 406-416. https://doi.org/10.1007/s004120050324