Efficiency of de novo centromere formation in human artificial chromosomes.
In a comparative study, we show that human artificial chromosome (HAC) vectors based on alpha-satellite (alphoid) DNA from chromosome 17 but not the Y chromosome regularly form HACs in HT1080 human cells. We constructed four structurally similar HAC vectors, two with chromosome 17 or Y alphoid DNA (17alpha, Yalpha) and two with 17alpha or Yalpha and the hypoxanthine guanine phosphoribosyltransferase locus (HPRT1). The 17alpha HAC vectors generated artificial minichromosomes in 32-79% of the HT1080 clones screened, compared with only approximately 4% for the Yalpha HAC vectors, indicating that Yalpha is inefficient at forming a de novo centromere. The 17alpha HAC vectors produced megabase-sized, circular HACs containing multiple copies of alphoid fragments (60-250 kb) interspersed with either vector or HPRT1 DNA. The 17alpha-HPRT1 HACs were less stable than those with 17alpha only, and these results may influence the design of new HAC gene transfer vectors.
Mejía
José E
JE
Weatherall Institute of Molecular Medicine, University of Oxford, Oxford, OX3 7BN, UK.
Alazami
Anas
A
Willmott
Adrian
A
Marschall
Peter
P
Levy
Elaine
E
Earnshaw
William C
WC
Larin
Zoia
Z
D002460:Cell Line; D002503:Centromere / Q000378:metabolism*; D022222:Chromosomes, Artificial, Human / Q000378:metabolism*; D002886:Chromosomes, Human, Pair 17 / Q000235:genetics*; D005822:Genetic Vectors; D006801:Humans; D007041:Hypoxanthine Phosphoribosyltransferase / Q000235:genetics; D014998:Y Chromosome / Q000235:genetics*
D007041:Hypoxanthine Phosphoribosyltransferase