Gene Targeting to ALU in Cultured Human Somatic Cells

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Cheryl Knox


Gene therapy holds the potential to cure many human genetic disorders. Gene targeting, the directed modification of chromosomal DNA via homologous recombination with an introduced plasmid vector, is one technique used in gene therapy. We hypothesized that targeting in ALU sequence, found interspersed throughout the genome at a high copy number, would improve the frequency of homologous recombination. To test this hypothesis, human somatic cells were transfected with the SLU containing vector pBP47. Recovery of integrated plasmids from the human genomic DNA will indicate the frequency of homologous recombination. Experiments to maximize plasmid recovery were performed. The specific human genomic DNA used for rescue in this study showed strong inhibition at the transformation stage of the rescue. Comparisons between individual genomic DNA preparations (human verses barley DNA) suggested that rescue inhibition was caused by either the human genomic DNA itself or something associated with it.