Human Molecular Genetics, Vol 7, 1635-1640, Copyright © 1998 by Oxford University Press
B Grimes and H Cooke
Construction of a mammalian artificial chromosome (MAC) will develop our
understanding of the requirements for normal chromosome maintenance,
replication and segregation while offering the capacity for introducing
genes into cells. Construction of MACs with telomere, centromere and
replication function has been approached by two methods. The 'top down'
strategy uses artificially induced chromosome truncations as a means to
define a minimal chromosome that retains the mitotic properties of a normal
chromosome. The 'build up' approach has focused on attempts to assemble MAC
vectors containing functionally defined telomere repeats together with
candidate centromere and replication origin sequences. Here we report on
significant advances in both areas, with particular emphasis on two reports
showing that stable, low copy number MACs containing a functional
centromere can be produced following transfection of naked DNA into the
human HT1080 cell line. One approach used a transfection mixture of cloned
synthetic alpha-satellite arrays up to 1 Mb in length and unlinked
telomeric DNA, in either the presence or absence of random human genomic
DNA fragments. In the second approach, MACs were formed from a defined
yeast artificial chromosome (YAC) DNA molecule containing 100 kb of highly
homo- geneous alphoid DNA retrofitted with human telomere repeats. These
results demonstrate for the first time that alpha- satellite DNA can seed
de novo centromeres in human cells, indicating that this repetitive
sequence family plays an important role in centromere function. The
stability of these MACs suggests that they have potential to be developed
as gene delivery vectors.
REVIEWS
Engineering mammalian chromosomes
MRC Human Genetics Unit, Western General Hospital, Crewe Road, Edinburgh EH4 2XU, UK. brenda@hgu.mrc.ac.uk
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