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        Poxviral/Retroviral Chimeric Vectors Allow Cytoplasmic Production of Transducing Defective Retroviral Particles

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        Retroviral vectors have become important tools in gene therapy as a result of a number of highly desirable properties, including stable genomic integration, which are rarely found with other gene delivery vectors. However, after two decades of experience with this technology, some shortcomings still remain to be solved. Retroviral vectors cannot be grown to as high titers, as for example, adenoviruses; they tend to be unstable and are sensitive to lysis by complement when transfused into patients. The search for more robust retroviral delivery systems has led to the development of hybrid viral vectors to combine the broadly estimated features of retroviral vectors with advantageous properties of another viral vector system. Chimeric systems with adeno- or alphavirus vectors have already been reported (1 3 ). Furthermore, herpesvirus amplicons expressing gag, pol , and env genes were shown to rescue defective lac Z retro virus in a cell line carrying a corresponding pro virus (4 ). This chapter is dedicated to vaccinia virus, one of the most widely used vectors in molecular and cell biology (5 ,6 ) as a chimeric carrier for small RNA viruses, such as retroviruses. The methods provided in this chapter disclose the necessary steps for the generation of such hybrid vectors and for their use to produce retroviral vector particles in packaging cells. However, the system offers a number of interesting opportunities that will require individual cloning concepts. To fully exploit the possibilities of the hybrid vector system, it is therefore advisable to consider the life cycle and the specific properties of the two vectors that are fused in this chimera.
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