Adeno x purification kit

The p32 plasmid, pCMV6-XL5, was purchased from OriGene Technologies, Inc. and subcloned into the BglII and KpnI restriction sites of the pCMV-Tag1 plasmid. For virus generation, full-length p32 was cloned into the BamHI and XhoI sites of the pENTR-CMV vector that has attL sites for site-specific recombination using the Gateway Destination Vector system (Invitrogen; Thermo Fisher Scienfitic, Inc.). The site-specific recombination between the pENTR-CMV/p32 and the adenovirus vector, pAd/PL-DEST, was conducted using LR clonase II. WT Adp32 is an adenovirus encoding full-length human p32. The adenovirus was amplified in 293A cells and purified using an Adeno-X™ purification kit (Takara Bio, Inc.) and the multiplicity of infection was determined using an Adeno-X™ titer kit (Takara Bio, Inc.). Adp32 was used to treat HUVECs at a concentration of 1×10⁶ pfu/ml. For in vivo mice experiments, the purified recombinant adenovirus containing 5×10⁹ particles was injected in the tail vein of mice. Adenovirus only as an empty vector (Ad) was used as an adenoviral control.