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        A Method for Helper Virus-Free Production of Adeno-Associated Virus Vectors

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        The need for safe and effective gene transfer to mammalian cells and organisms has led many investigators to focus on the defective human parvovirus, adeno-associated virus (AAV), as a gene delivery vehicle. AAV vectors have been used to deliver genes to a wide variety of mammalian cells in culture, as well as to brain (1 ), retina (2 ,3 ), cochlea (4 ), skeletal muscle (5 ,6 ), cardiac muscle (7 ), liver (8 ), lung (9 ,10 ), central nervous system (11 ), and skin (12 ). The success of these preclinical models has led to several ongoing and proposed human clinical trials (13 ). One of the major limitations of the AAV vector system to date has been the lack of a versatile and robust method of vector preparation. This chapter describes a method for the helper virus-free preparation of AAV vectors. The method circumvents the need for adenovirus (Ad) coinfection and its simplicity makes it amenable for screening multiple AAV vector constructions. To lay the foundation for the development of this and other AAV packaging systems, a brief summary of AAV biology is provided later. More detailed reviews describing AAV and its derived vectors are available in (14 and 15 ).
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