Construction of replication-defective herpes simplex virus vectors.

Advances in identification and characterization of gene products responsible for specific diseases of the nervous system have opened opportunities for novel therapies using gene transfer vectors for gene replacement. Herpes simplex virus (HSV)-based vectors are particularly well suited for gene delivery to neurons of the central and peripheral nervous systems. The authors have developed methods to delete HSV-1 IE gene functions and to subsequently introduce foreign genes into the HSV-1 genome using homologous recombination. This unit describes methods for generating cell lines that complement multiple essential gene deletion mutants as well for generating such replication-defective virus recombinants and inserting foreign DNA sequences into replication-defective viral genomes, the last step in preparing a vector. Three support protocols describe methods for preparing virus stocks, titering virus, and preparing viral DNA.