Advantages and disadvantages of multiple different methods of adenoviral vector construction

Adenoviruses are medium-sized nonenveloped DNA tumor viruses whose genome consists of a linear double-stranded DNA molecule of about 36 Kb (reviewed in refs. 1-3). Adenoviruses have icosahedral geometry with a diameter of approx 70-100 nm. Currently, over 100 different mammalian and foul adenovirus (Ad) serotypes have been characterized, with 49 of them being human serotypes. The most extensively characterized serotypes are 2, 5, and 12, with type-5-based vectors being the most popular. Ad virus transcription, replication, and packaging have been extensively studied and take place in the nucleus of infected cells, with transcription and replication requiring both cellular and viral proteins. Expression of the Ad genome is divided into two phases: early, which occurs before DNA replication, and late, which occurs after the initiation of DNA replication. Transcription during the early phase of adenoviral infection involves the highly orchestrated expression of four pol IIdependent regions, E1, E2, E3, and E4, which each code for multiple mRNAs. E1 is further divided into two transcriptional units E1A and E1B. E1A coordinates transcriptional control of the virus, whereas E1B gene products influence the early to late transition by coordinating viral to cellular mRNA metabolism and host protein shutoff. Additionally, the E1B region obstructs the apoptotic signals induced by E1A. The E2 region codes for three proteins essential for viral DNA replication; terminal protein precursor, DNA polymerase, and DNA.