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Epstein-Barr-based episomal chromosomes shuttle 100 kb of self-replicating circular human DNA in mouse cells
Z T Kelleher1, H Fu, E Livanos
1Lineberger Comprehensive Cancer Center, University of North Carolina at Chapel Hill 27599, USA.
Abstract:
We describe the microcell fusion transfer of 100-200 kb self-replicating circular human minichromosomes from human into mouse cells. This experimental approach is illustrated through the shutting of the latent 170 kb double-stranded DNA genome from the human herpesvirus, Epstein-Barr virus, into nonpermissive rodent cells. Using this interspecies transfer strategy, circular episomes carrying 95-105 kb of human DNA were successfully established at low copy number in mouse A9 cells. Selected episomes were stably maintained for 6 months, and unselected episomes were characterized by a 95% episomal retention per cell division. The establishment of a mouse artificial episomal chromosome system should facilitate evolutionary and therapeutic studies of large human DNA in rodent genetic backgrounds.
Insights
Researchers transferred large human DNA minichromosomes into mouse cells, achieving stable maintenance and high retention rates. This breakthrough enables new studies of human DNA in rodent models.
Area of Science:
- * Molecular Biology
- * Genetics
- * Cell Biology
Background:
- * Transferring large DNA fragments between species presents significant technical challenges.
- * Human minichromosomes offer a way to study large DNA elements in a manageable format.
Purpose of the Study:
- * To establish a method for transferring and maintaining large, self-replicating human DNA minichromosomes in mouse cells.
- * To demonstrate the feasibility of interspecies transfer for creating artificial episomal chromosome systems.
Main Methods:
- * Microcell-mediated chromosome transfer was employed to introduce human minichromosomes into mouse A9 cells.
- * The Epstein-Barr virus (EBV) latent genome served as a model for demonstrating the transfer of a 170 kb double-stranded DNA episome.
- * Interspecies transfer strategy utilized circular episomes carrying 95-105 kb of human DNA.
Main Results:
- * Successfully established circular episomes with 95-105 kb of human DNA in mouse A9 cells at a low copy number.
- * Demonstrated stable maintenance of selected episomes for up to 6 months.
- * Achieved high episomal retention (95% per cell division) for unselected episomes.
Conclusions:
- * The successful interspecies transfer and stable maintenance of human minichromosomes in mouse cells create a novel artificial episomal chromosome system.
- * This system provides a powerful platform for evolutionary studies of large human DNA.
- * Facilitates therapeutic research involving large human DNA fragments in rodent genetic backgrounds.