Fifty-One Kilobase HSV-1 Plasmid Vector Can Be Packaged Using a Helper Virus-Free System and Supports Expression in the Rat Brain

Herpes simplex virus type 1 (HSV-1) plasmid vectors have a number of attractive features for gene transfer into neurons. In particular, the large size of the HSV-1 genome suggests that HSV-1 vectors might be designed to accommodate large inserts. We now report the construction and characterization of a 51 kb HSV-1 plasmid vector. This vector was efficiently packaged into HSV-1 particles using a helper virus-free packaging system. The structure of the packaged vector DNA was verified by both Southern blot and PCR analyses. A vector stock was microinjected into the rat striatum, the rats were sacrificed at 4 days after gene transfer, and numerous X-gal positive striatal cells were observed. This 51 kb vector was constructed using general principles that may support the routine construction of large vectors. Potential applications of such HSV-1 vectors include characterizing large promoter fragments or genomic clones and co-expressing multiple genes.