Bi-HAC vector system toward gene and cell therapy.

Genetic manipulations with mammalian cells often require introduction of two or more genes that have to be in trans-configuration. However, conventional gene delivery vectors have several limitations, including a limited cloning capacity and a risk of insertional mutagenesis. In this paper, we describe a novel gene expression system that consists of two differently marked HAC vectors containing unique gene loading sites. One HAC, 21HAC, is stably propagated during cell divisions; therefore, it is suitable for complementation of a gene deficiency. The other HAC, tet-O HAC, can be eliminated, providing a unique opportunity for transient gene expression (e.g., for cell reprogramming). Efficiency and accuracy of a novel bi-HAC vector system have been evaluated after loading of two different transgenes into these HACs. Based on analysis of transgenes expression and HACs stability in the proof of principle experiments, the combination of two HAC vectors may provide a powerful tool toward gene and cell therapy.