I did AAV packaging more than 10 times, and every time went well. But recently, the titer dropped greatly. I checked cell status and changed more healthy and younger cells, and I also re-purified plasmids and ran gel to test their size and supercoil. But no matter I used PDG or RC and Helper, both titers were low. The strangest thing is that the transfection rate was 70% up, and aav-gfp was quite bright. While my real-time PCR and infection experiment showed the titer was not ideal at all.
I used PEG-8000 to condense viruses and it was feasible in previous experiment. The AAV system is from Stratagene, and have a GFP marker in expression plasmid.
I cannot think out more factors which may influence aav packaging. Could anybody help me?
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