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Manipulation of human minichromosomes to carry greater than megabase-sized chromosome inserts

Nature Biotechnology volume 18, pages 1086–1090 (2000)Cite this article

Abstract

For introducing regions of human chromosomes greater than a megabase into cells or animals, we have developed a chromosome-cloning system in which defined regions of human chromosomes can be cloned into a stable human minichromosome vector in homologous recombination-proficient chicken DT40 cells. The stable minichromosome vector allowed a 10 Mb-sized region of the mitotically unstable human chromosome 22 to be stably maintained in mouse embryonic stem (ES) cells, and in mice. Furthermore, we demonstrated functional expression of human genes from the HAC in mice. This study describes a stable cloning and expression system for greater than megabase–sized regions of human chromosomes.

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Figure 1: Schematic diagram of the chromosome-cloning approach.
Figure 2: Cloning of the hCI22 and hCI2 into SC20 vector.
Figure 3: Verification of successful isolation of the cells containing λ-HAC.
Figure 4: Mitotic stability of λ-HAC in mouse ES cells.
Figure 5: The function of λ-HAC in chimeric mice.

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Acknowledgements

We thank Dr. Shun-ichi Takeda for technical advice and valuable discussion; Saori Suzuki and Sumiko Igami for technical assistance.

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Author notes

  1. Yoshimi Kuroiwa and Kazuma Tomizuka: These authors contributed equally to this work.

Authors and Affiliations

  1. Pharmaceutical Research Laboratory, Kirin Brewery. Co., Ltd., 3 Miyahara-cho, Takasaki-shi, 370-1295, Gunma, Japan

    Yoshimi Kuroiwa, Kazuma Tomizuka, Hitoshi Yoshida, Atsuko Ohguma, Terumi Yamamoto, Sonoko Tanaka & Isao Ishida

  2. Department of Molecular and Cell Genetics, School of Life Sciences, Faculty of Medicine, Tottori University, Nishimachi 86 Yonago, Tottori, 683-8503, Japan

    Tokuyuki Shinohara, Yasuhiro Kazuki & Mitsuo Oshimura

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  1. Yoshimi Kuroiwa
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Correspondence to Isao Ishida.

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Kuroiwa, Y., Tomizuka, K., Shinohara, T. et al. Manipulation of human minichromosomes to carry greater than megabase-sized chromosome inserts. Nat Biotechnol 18, 1086–1090 (2000). https://doi.org/10.1038/80287

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