A recombinant E1-deleted porcine adenovirus-3 as an expression vector

Abstract

Replication-defective E1-deleted porcine adenoviruses (PAVs) are attractive vectors for vaccination. As a prerequisite for generating PAV-3 vectors containing complete deletion of E1, we transfected VIDO R1 cells (fetal porcine retina cells transformed with E1 region of human adenovirus 5) with a construct containing PAV-3 E1Blarge coding sequences under the control of HCMV promoter. A cell line named VR1BL could be isolated that expressed E1Blarge of PAV-3 and also complemented PAV214 (E1A+E1Bsmall deleted). The VR1BL cells could be efficiently transfected with DNA and allowed the rescue and propagation of recombinant PAV507 containing a triple stop codon inserted in the E1Blarge coding sequence. In addition, recombinant PAV227 containing complete deletion of E1 (E1A+E1Bsmall + E1Blarge) could be successfully rescued using VR1BL cell line. Recombinant PAV227 replicated as efficiently as wild-type in VR1BL cells but not in VIDO R1 cells, suggesting that E1Blarge was essential for replication of PAV-3. Next, we constructed recombinant PAV219 by inserting green fluorescent (GFP) protein gene flanked by a promoter and a poly(A) in the E1 region of the PAV227 genome. We demonstrated that PAV219 was able to transduce and direct expression of GFP in some human cell lines.