Cre-dependent viral vector for optogenetics
We have developed a widely used Cre-dependent viral vector utilizing a genetic element called a FLEX switch.
Plasmids are available from Addgene.com
These vectors are prone to recombination. This is a well known issue with these AAV vectors and is due to the inverted terminal repeats (ITRs) required for rAAV production. To minimize recombination, we propagate these plasmids in Stbl2 cells from Invitrogen. Also, to minimize recombination, cells should be cultured at 30 ºC.
Note that these cultures will grow slowly (20 h for minipreps). Better yields and culture times are obtained with 2xYT as the media. This is strongly recommended.
Because recombination may still happen occasionally, we do a panel of restriction digestions to assess whether the ITRs are in tact. Separate digestions with PvuII, Sma1, and SnaB1 should be performed. The expected patterns can be calculated from the attached sequence available on addgene.com.
Viruses for Cre-dependent optogentics based on FLEX switch available from University of Pennsylvania Vector Core.