Inverted terminal repeats of adeno-associated virus decrease random integration of a gene targeting fragment in Saccharomyces cerevisiae

Table 2 Effect of the AAV Rep protein expression on gene targeting and random integration in yeast

Plasmid	Gene targeting	Random integration	GT/RI
ITR-lys2URA3lys2-ITR, No REP	31.8 ± 9.8	0.73 ± 0.39	43.5
ITR-lys2URA3lys2-ITR, REP	20.1 ± 5.9	0.88 ± 0.49	22.8
ITR-pokURA3-ITR, No REP	ND	0.12 ± 0.08	/
ITR-pokURA3-ITR, REP	ND	1.02 ± 0.56*	/

Yeast was transformed with Pvu II-restricted pAAVLUL or pAAVpokURA as reported in Materials and Methods. In the case of experiments with pAAVLUL, gene targeting and random integration events are reported as number of URA3⁺lys2^-colonies /10³ transformants per μg of plasmid DNA and URA3⁺LYS2⁺ colonies /10³ transformants per μg of plasmid DNA, respectively. Similarly, in the case of pAAVpokURA experiments, non-homologous integration events are reported as total URA3⁺colonies /10³ transformants per μg DNA. The number of transformants per μg of plasmid DNA was determined by transformation with 1 μg of episomal plasmid DNA. Results are the mean of four independent experiments ± standard deviation. The number of total colonies is indicated in the parentheses. Statistics was performed using Student’s t-test. *p = 0.0078 vs ITR-pokURA3-ITR, No REP. ND = not detectable.

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