Utilization of non-viral sequences for minus-strand DNA transfer and gene reconstitution

摘要

A retroviral vector for gene reconstitution is provided that includes a 3' portion of a heterologous nucleic acid sequence 5' of a first att site of the retroviral vector, a 5' portion of the heterologous nucleic acid sequence 3' of a second att site of the retroviral vector. In this vector, a sub-portion of the 3' portion of the heterologous nucleic acid sequence and the 5' region of the heterologous nucleic acid sequence are direct repeats. Transformation of a eukaryotic cell with the retroviral vector results in reconstitution and duplication of the heterologous nucleic acid sequence. A retroviral vector for gene reconstitution is also provided that includes a 3' portion of a heterologous nucleic acid sequence inserted into or adjacent to a 5' retroviral terminal repeat of the retroviral vector, and a 5' portion of the heterologous nucleic acid sequence inserted into or adjacent to a 3' retroviral terminal repeat of the retroviral vector, wherein the 5' retroviral terminal repeat and the 3' retroviral terminal repeat each comprise an att site. A sub-portion of the 3' portion of the heterologous nucleic acid sequence and the 5' region of the heterologous nucleic acid sequence are direct repeats. Transformation of a eukaryotic cell with this retroviral vector results in reconstitution and duplication of the heterologous nucleic acid sequence. Method and kits are provided for reconstituting and duplicating a nucleic acid molecule in a host cell using these retroviral vectors to introduce a heterologous nucleic acid in the cell. A method is provided for treating a subject that includes contacting a cell of the subject with a therapeutically effective amount of a retroviral vector for gene reconstitution and duplication.