Stable (Heritable) DNA Transfer without Antibiotic Selection?

On Mendiburo et al., 2011 in Science

     Centromeres are the chromatin regions that are specialized to carry out the chromosome segregation during cell division. Because the spindle attachment sites are stable and inherited, initial analyses suggested that centromeres were sequence specific and the centromere formation is dictated by DNA sequence alone.  However, in very rare occasions centromeres can leave this stable spot and move to a new chromosomal site. These "neocentromeres" are as valid as natural centromeres and segregate the chromosomes with high fidelity.  This finding implies that the source of the specificity and the heritability of the centromeres might not be genetic but rather epigenetic.
A clue comes from the finding that specialized histone H3, called centromere protein A (CENP-A), is incorporated to the nucleosomes at centromeric loci. Mendiburo et al, hypothesized that this special histone could mark the centromeres on the chromosome. (Mendiburo et al., 2012) To test this they fused  CENP-A to bacterial LacI protein and engineered a LacO site  (which binds to the LacI) in their cell culture model. Surprisingly CENP-A was localized to the LacO and was incorporated to the local nucleosomes and recruited even the endogenous CENP-A to the new site.  This neocentromere was inherited and associated with spindle assembly normally. Although adding a second centromere causes mitotic failure and cell death, they also show that plasmids engineered like this can act as artificial chromosomes.
This brings up the idea that recombinant DNA transfected to eukaryotic cultured cells can be made heritable and stable without the need for heavy antibiotic selection and genome integration. Antibiotic selection is very strong and kills all the cells without the plasmid and the cells which looses the plasmid and only genome integrated copies remain. Some cells are fragile and do not survive this process. Another disadvantage is that genome integration is random and might cause aberrations if the integration site is vital. This is especially important in the case of gene therapy because genome integration could cause additional defects while mending the disease.

María José Mendiburo, 

Jan Padeken, Stefanie Fülöp,Aloys Schepers and Patrick HeunDrosophila CENH3 Is Sufficient for Centromere FormationScience 4 November 2011: 334 (6056), 686-690. [DOI:10.1126/science.1206880]