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Human Molecular Genetics, 2000, Vol. 9, No. 12 1891-1902
© 2000 Oxford University Press

Human mini-chromosomes with minimal centromeres

J.W. Yang+, C. Pendon§, J. Yang, N. Haywood, A. Chand{dagger} and W.R.A. Brown{ddagger}

Biochemistry Department, South Parks Road, Oxford OX1 3QU, UK

We have introduced a 6.5 Mb human mini-chromosome with a complex centromere structure into DT40 cells and have used sequence targeting and telomere-directed chromosome breakage to dissect the sequence requirements for centromere function. These experiments proved that a vertebrate centromere with two blocks of functional alphoid DNA separated by 2.5 Mb can exist as a stable structure in some but not all vertebrate cells. Further experiments indicated that recovery of chromosomes with less than ~100 kb of alphoid DNA is very inefficient, suggesting that a functional centromere requires a minimum of ~100 kb of alphoid DNA. Mini-chromosomes with minimal centromeres segregate accurately in some but not all vertebrate cells and should be useful for the detection of sequence-specific factors required for vertebrate centromere maintenance.

+ Present address: MRC Mammalian Genetics Unit, Harwell, Oxfordshire OX11 ORD, UK§Present address: Laboratorio de Bioquimica y Biologia Molecular, Facultad de Ciencias, Apdo 40, 11510 Puerto Real, Cadiz, Spain ¶Present address: Wellcome/CRC Institute, Tennis Court Road, Cambridge CB2 1QR, UK {dagger}Present address: ISIS Innovation, Ewert House, Summertown, Oxford OX2 7BZ, UK{ddagger}To whom correspondence should be addressed at: Institute of Genetics, University of Nottingham, Queen’s Medical Centre, Nottingham NG7 2UH, UK. Tel: +44 115 849 3244; Fax: +44 115 970 9906; Email: william.r.brown@nottingham.ac.uk


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