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Nucleic Acids Research, 1988, Vol. 16, No. 14 7103-7117
© 1988

Articles

Reconstruction of the yeast 2 µm plasmid partitioning mechanism

M.J. Dobson*, F.E. Yull1, M. Molina2, S.M. Kingsman1 and A.J. Kingsman1

Department of Botany, University of Nottingham University Park, Nottingham, NG7 2RD 1Department of Biochemistry South Parks Road, Oxford, OX1 3QU, UK 2Department de Microbiologia, Facultad de Farmicia, Universidad Complutense, Ciudad Universitaria Madrid-3, Spain

* To whom correspondence should be addressed.

Received May 20, 1988. Revised June 22, 1988. Accepted June 22, 1988.

The effect of the yeast 2µm circle encoded REP1 and REP2 gene products on plasmid partitioning and copy number control was analyzed by removing the open reading frames from their normal sequence context and transcriptional control regions and directing their expression using heterologous promoters in [cir0] host strains. Both the REP1 and REP2 gene products are directly required at appropriate levels of expression to reconstitute the 2µm circle partitioning system in conjunction with REP3 and the origin of replication. The level of expression of REP2 appears to be critical to re-establishing proper partitioning and may also play a role in monitoring and thereby regulating the plasmid copy number. Constitutive expression of the REP1 and REP2 open reading frames using heterologous expression signals can be used to reconstruct efficient plasmid partitioning even in the absence of FLP-mediated plasmid amplification and a functional D open reading frame.


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