Reversibility and adjustability of macrolide-responsive SEAP expression in HT-1080 and MCF-7 transduced by transgenic AAV type 2-derived particles

<p><b>Copyright information:</b></p><p>Taken from "Adeno-associated viral vectors engineered for macrolide-adjustable transgene expression In mammalian cells and mice"</p><p>http://www.biomedcentral.com/1472-6750/7/75</p><p>BMC Biotechnology 2007;7():75-75.</p><p>Published online 6 Nov 2007</p><p>PMCID:PMC2211474.</p><p></p> (A) HT-1080 and (B) MCF-7 were transduced with pDF143-derived AAV particles (2000 genomic particles/cell) and six equal populations (1–6) were cultivated in media containing different antibiotic concentrations. Cells were (i) cultivated in the presence (+++) or absence (---) of EM over 9 days, (ii) cultivated in the presence (++-) or absence (--+) of EM over 6 days and then incubated in reversed EM conditions for the remaining three days or (iii) cells were cultivated in medium whose EM status was alternated every three days +EM to --EM to +EM (+-+) or from --EM to +EM to --EM (-+-) and SEAP expression was quantified. Adjustability of SEAP expression of pDF143 (1000 genomic particles/cell) (C) and pDF51/77 (500 genomic particles/cell) (D) in HT-1080 and MCF-7 cultivated for 48 h in the presence of increasing EM concentrations. SEAP expression is shown in units/liter (U/l) as defined by Schlatter et al. [52]. Abbreviations: EM, erythromycin; SEAP, human placental secreted alkaline phosphatase. Abbreviations: EM, erythromycin; SEAP, human placental secreted alkaline phosphatase

Macrolide-triggered SEAP expression in mice injected with pDF143 (ITR-P-ET1-pA-P-SEAP-pA-ITR)-derived AAV type 2 particles

<p><b>Copyright information:</b></p><p>Taken from "Adeno-associated viral vectors engineered for macrolide-adjustable transgene expression In mammalian cells and mice"</p><p>http://www.biomedcentral.com/1472-6750/7/75</p><p>BMC Biotechnology 2007;7():75-75.</p><p>Published online 6 Nov 2007</p><p>PMCID:PMC2211474.</p><p></p> pDF143-derived AAV particles were administered intramuscularly and SEAP levels in the serum were measured at two different time points for one group in the absence of erythromycin, and for the other group with EM injections every 24 h. SEAP expression is shown in milliunits/liter (mU/l) as defined by Schlatter et al. [52]. Abbreviations: EM, erythromycin; SEAP, human placental secreted alkaline phosphatase

Self- and auto-regulated AAV type 2-based expression of fluorescent proteins

<p><b>Copyright information:</b></p><p>Taken from "Adeno-associated viral vectors engineered for macrolide-adjustable transgene expression In mammalian cells and mice"</p><p>http://www.biomedcentral.com/1472-6750/7/75</p><p>BMC Biotechnology 2007;7():75-75.</p><p>Published online 6 Nov 2007</p><p>PMCID:PMC2211474.</p><p></p> (A) Schematic representation of an auto-regulated (pDF124), a bidirectional (pDF89) and a self-regulated (pDF141) AAV type 2-based expression unit. (B) Fluorescence micrographs of human fibrosarcoma cells (HT-1080) and a human breast cancer cell line (MCF-7) transduced with pDF124-, pDF89- and pDF141-derived AAV particles (2000 genomic particles/cell) cultivated in the presence (+) and absence (-) of EM. (C) FACS-mediated quantification of EYFP in HT-1080 transduced with equal amounts of viral particles (2000 genomic particles/cell) and cultivated in the presence (+EM) and absence (-EM) of erythromycin. Abbreviations: EM, erythromycin; ET1, erythromycin-dependent transactivator; ETR, ET1-specific operator; EYFP, enhanced yellow fluorescent protein; IRES, internal ribosome entry site; ITR, inverted terminal repeat; pA, synthetic polyadenylation signal; pA, simian virus 40 polyadenylation signal; P, erythromycin-responsive promoter; P, minimal version of the heat-shock protein 70 promoter; P, simian virus 40 promoter