Ordering
| Item | Catalog # | Description | Quantity | Price (USD) | ||
|---|---|---|---|---|---|---|
| Plasmid | 68720 | Standard format: Plasmid sent in bacteria as agar stab | 1 | $75 | Add to Cart | |
| AAV1 | 68720-AAV1 | Virus (100 µL at titer ≥ 1×10¹³ vg/mL)and Plasmid.More Information | Add to Cart | |||
This material is available to academics and nonprofits only.
Backbone
- Vector backbonepAAV(Search Vector Database)
- Total vector size (bp)6958
- Vector typeAAV
Growth in Bacteria
- Bacterial Resistance(s)Ampicillin
- Growth Temperature37°C
- Growth Strain(s)Stbl3
- Growth instructionsDue to ease of recombination, AAV and lentivirus vectors should be amplified in a recombination deficient bacteria strain such as Invitrogen's OneShot Stbl3 cells. Check for integrety of ITR sites with SmaI digest.
- Copy numberLow Copy
Gene/Insert
- Gene/Insert namemRuby2-P2A-GCaMP6s
- SpeciesR. norvegicus (rat), G. gallus (chicken); A. victoria (jellyfish)
- Insert Size (bp)2124
- MutationCre-dependent expression from inverted open reading frame (i.e. 'FLEXED')
- PromoterhSyn1-FLEX
Cloning Information
- Cloning methodUnknown
- 5′ sequencing primeractcagcgctgcctcagtct
- 3′ sequencing primergtttgtacaaatgatgacagcgaag (Common Sequencing Primers)
Resource Information
- Terms and Licenses
- UBMTA
- Ancillary Agreement for Plasmids Containing FP Materials
- genOway Notice of RIghts
- Industry Terms
- Not Available to Industry
- Article Citing this Plasmid
- 1 Reference
Depositor Comments
mRuby2: Article: Improving FRET dynamic range with bright green and red fluorescent proteins. Lam et al (Nat Methods. 2012 Sep 9. doi: 10.1038/nmeth.2171. PubMed) Addgene Plasmid 40260
GCaMP6s: Article: Ultrasensitive fluorescent proteins for imaging neuronal activity. Chen et al (Nature. 2013 Jul 18;499(7458):295-300. doi: 10.1038/nature12354. PubMed)Addgene Plasmid 40753
Information for AAV1 (Catalog # 68720-AAV1)(Back to top)
Purpose
Ready-to-use AAV1 particles produced from pAAV-hSyn1-Flex-mRuby2-GSG-P2A-GCaMP6s-WPRE-pA (#68720). In addition to the viral particles, you will also receive purified pAAV-hSyn1-Flex-mRuby2-GSG-P2A-GCaMP6s-WPRE-pA plasmid DNA.
Cre-dependent GCaMP6s calcium sensor and bicistronic, physically separate mRuby2 expression under a human synapsin1 promoter. These AAV preparations are suitable purity for injection into animals.Delivery
- Volume100 µL
- Titer≥ 1×10¹³ vg/mL
- Pricing$350 USD for preparation of 100 µL virus + $30 USD for plasmid.
- StorageStore at -80℃. Thaw just before use and keep on ice.
- ShipmentViral particles are shipped frozen on dry ice. Plasmid DNA (≥ 200ng) will also be included in the shipment.
Viral Production & Use
- Packaging Plasmidsencode adenoviral helper sequences and AAV rep gene, AAV1 cap gene
- BufferPBS + 0.001% Pluronic F-68
- SerotypeAAV1
- PurificationIodixanol gradient ultracentrifugation
- Reporter GenemRuby2 (Cre-dependent)
Biosafety
Requestor is responsible for compliance withtheir institution"s biosafety regulations.Lentivirus is generally considered BSL-2. AAV isgenerally considered BSL-1, but may requireBSL-2 handling depending on the insert.Biosafety Guide
Resource Information
- Terms and Licenses
- Ancillary Agreement for Penn Vectors
- Terms of Use for Viral Vectors
- Industry Terms
- Not Available to Industry
Viral Quality Control
- Addgene ensures high quality viral vectors by optimizing and standardizing production protocols and performing rigorous quality control (QC) (see a list of our QC assays). Thespecific QC assays performed varies for each viral lot. To learn which specific QC assays were performed on your lot, please contact us.
- Titer: the exact titer of your sample will be reported on the tube. The titer you see listed on this page is the guaranteed minimum titer. See how titers are measured.
Visit our viral production page for moreinformation.
Addgene Comments
Using FLEX vectors in vivo: LoxP sites in FLEX plasmids are known to recombine during DNA amplification and viral vector production, which may result in a minority of Cre-activated (i.e., "flipped") viral vectors. Addgene has measured this occurs in 0.01-0.03% of viral vectors in our typical production protocol. This can lead to a small number of cells exhibiting Cre-independent transgene expression in vivo. To address this, we recommend titrating to find the optimal AAV dosage required for Cre-dependent transgene expression and function in vivo. This may include reducing the viral vector dosage in order to reduce the likelihood of Cre-independent expression.
