Recombinant vectors based on adeno-associated virus (AAV) or human immunodeficiency 1 (lentivirus) are promising tools for long term in vivo gene delivery. Their design allows the exchange of capsids or envelopes, respectively, theoretically providing the opportunity to transduce a range of cell types. We constructed AAV vectors encoding enhanced green fluorescent protein (EGFP) within an AAV serotype 2 (AAV2) genome contained in an AAV2, five or one capsid (called AAV2/2, AAV2/5 and AAV2/1, respectively). Similarly we produced lentiviral vectors, encoding the same expression cassette present in the AAV vectors, pseudotyped with proteins from vesicular stomatitis virus glycoprotein (VSVG) or Mokola envelopes. Transduction characteristics of these vectors were evaluated in the murine retina following subretinal or intravitreal administration. The time of onset of transgene expression and the targeted cell types differed between the various recombinants. Onset of transgene expression was 3-4 days for lentiviral vectors and AAV2/1. In contrast, onset was at 2-4 weeks for AAV2/5 and AAV2/2, respectively. After subretinal injection, both lenti-VSVG and AAV2/5 transduced the retinal pigment epithelium (RPE) and photoreceptors efficiently whereas transgene expression was restricted to RPE cells using lenti with the Mokola envelope or AAV2/1. After intravitreal administration, only AAV2/2 and lenti-VSVG transduced the inner retina. Vector-mediated fluorescence was detected in the retina for over 12 weeks for all of the vectors. We conclude that pseudotyping provides a useful means to manipulate viral vector cell targeting specificity as well as retinal transduction characteristics of vectors containing the same genome.