|Item||Catalog #||Description||Quantity||Price (USD)|
|Plasmid||30541||Plasmid sent as bacteria in agar stab||1||$65|
This material is available to academics and nonprofits only.
Backbone manufacturerAddgene plasmid 18917
- Backbone size w/o insert (bp) 5079
Vector typeAAV, Cre/Lox
Growth in Bacteria
Growth instructionsStbl2 E coli from Invitrogen Grow at 30 degrees in 2xYT media (see comments section below for more information). Please use 20 ug/ml of Amp.
Copy numberLow Copy
Insert Size (bp)1077
/ Fusion Protein
- tdTomato (C terminal on insert)
- Cloning method Restriction Enzyme
- 5′ cloning site KpnI (not destroyed)
- 3′ cloning site XbaI (not destroyed)
- 5′ sequencing primer pCAG-F
- 3′ sequencing primer WPRE-R (Common Sequencing Primers)
Please note this plasmid has a I170V in channelopsin 1. The changed amino acid improves the protein’s function.
To minimize recombination, propagate this plasmids in Stbl2 cells from Invitrogen. Also, to minimize recombination, cells should be cultured at 30 C.
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.
For more information on growth conditions, please see www.addgene.org/18917 as this is the backbone vector. 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. Please see Reviews in the right column for an image of Addgene's digest with these enzymes.
These plasmids were created by your colleagues. Please acknowledge the Principal Investigator, cite the article in which the plasmids were described, and include Addgene in the Materials and Methods of your future publications.
For your Materials & Methods section:AAV-Flex-rev-oChIEF-tdTomato was a gift from Rory McQuiston (Addgene plasmid # 30541)
For your References section:Nicotinic excitatory postsynaptic potentials in hippocampal CA1 interneurons are predominantly mediated by nicotinic receptors that contain alpha4 and beta2 subunits. Bell KA, Shim H, Chen CK, McQuiston AR. Neuropharmacology. 2011 Aug 25. 10.1016/j.neuropharm.2011.08.024 PubMed 21878344