Figure 1.

Design of Constructs and Confirmation of Infectivity

(A) The packaging construct (top) containing both the AAV2 Rep and Cap genes from AAV2, AAV8, or AAV9 were cloned into a pFastBac Dual vector. The Cap cassette is driven by the p10 promoter and contains the herpes simplex virus (HSV) thymidine kinase (tk) polyadenylation signal (pA). The Rep cassette is driven by the polyhedrin promoter (pH) and contains the simian virus 40 (SV40) pA. The transgene construct (bottom) contains either the ZsGreen or luciferase gene, which is driven by the cytomegalovirus (CMV) promoter. It also contains a beta-globin (B-globin) intron and a human growth hormone (hGH) pA signal. Lastly, the transgene cassette is flanked by the AAV2 inverted terminal repeat (ITR). (B) Purified AAV2 containing the ZsGreen transgene cassette was used to infect 293T cells at an MOI of 200. The cells were then imaged for ZsGreen expression 48 h postinfection. (C) Purified baculovirus containing the ZsGreen transgene cassette only was used to infect 293T cells at an MOI of 200. The cells were then imaged for ZsGreen expression 48 h postinfection.