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LINEAR PLASMIDS AND ARTIFICIAL CHROMOSOMES AS VECTORS FOR GENE TRANSFER IN EUKARYOTES

Objective


Research has been carried out to isolate and study replication origins and telomeres and to assemble the different elements into linear vectors, in order to improve gene transfer in eukaryotes. The replication origins studied came from viruses (bovine papilloma virus (BPV); simian virus 40 (SV40) and African swine fever virus (ASFV)) and chromosomes from animal cells. Plasmids containing the BPV origin of replication function could be injected into Xenopus embryos. Much is known about the structure of telomeres from lower eucaryotes and these sequences were used for the assembly experiments. In addition, telomeric segments were isolated and analyzed from human cells and ASFV. The role of simple repeating deoxyribonucleic acid (DNA) sequences on integration was determined and a putative signal for integration identified. Several linear vectors were assembled and their stability determined. Extracts from cells infected with ASFV enabled the in vitro replication of DNA. Human telomeric sequences were cloned in Escherichia coli and in yeast, and theviral telomeres of ASFV were isolated. A putative signal for integration (d(CG)) was identified. A linear vector containing the BVP origin of replication and Tetrahymena telomeres was shown to be stable in Xenopus oocytes.

Funding Scheme

CSC - Cost-sharing contracts

Coordinator

CONSEJO SUPERIOR DE INVESTIGACIONES CIENTIFICAS
Address
Serrano 117
Madrid
Spain