Cloning of the Adeno-Associated Virus Genome
The adeno-associated viruses (AAV) are defective parvoviruses that are dependent on coinfection with either adenovirus or herpesvirus for replication to occur (1–4). AAV has a linear single-strand DNA genome and is unusual in that strands of both polarities are encapsidated with equal frequency into separate virions (5,6). The nucleotide sequence of the entire genome of the human AAV2 has been determined and contains 4675 bases (7). There is an inverted terminal repetition of 145 bases, the first 125 bases of which form an overall palindromic sequence (Figure 1). Bases 1–41 are complementary to 125–85 while the internal sequence from 42–84 contains two shorter palindromes (42–62 and 64–84) (8). When the terminal palindromic sequence is folded on itself to maximize potential base-pairing, the T-shaped structure illustrated in Figure 1 is formed. In this structure only 7 of the terminal 125 bases are unpaired, 3 T’s and 3 A’s, the minimal number of bases required to be unpaired to allow the two short internal palindromic sequences to hairpin on themselves and a single T that separates the two internal palindromes. A second unusual feature of AAV is the existence of two alternative sequences from bases 62–84 that occur with equal frequency at both ends of the DNA (8,9). The alternative sequences result from an inversion of the terminal 125 bases during DNA replication as described below. If the sequence from 1–125 constituted a simple palindrome only the single odd base in the middle (base 63) would be affected by the inversion. In fact, however, the overall palindrome is interrupted by the two short internal palindromes from 42–84 and so this entire sequence is altered by inversion of the terminal 125 nucleotides.
KeywordsTerminal Repeat Terminal Sequence Inverted Terminal Repetition Palindromic Sequence Helper Virus
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