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pAAV-FLEX-ArchT-tdTomato
(Plasmid #28305)

Ordering

Item Catalog # Description Quantity Price (USD)
Plasmid 28305 Plasmid sent as bacteria in agar stab 1 $65
AAV5 28305-AAV5 Virus (100 µL at titer ≥ 7×10¹² vg/mL)
and Plasmid. More Information
$380

This material is available to academics and nonprofits only.

Backbone

  • Vector backbone
    AAV-FLEX
  • Backbone manufacturer
    Scott Sternson
  • Backbone size w/o insert (bp) 6482
  • Vector type
    Mammalian Expression, AAV, Cre/Lox ; Adeno-associated virus

Growth in Bacteria

  • Bacterial Resistance(s)
    Ampicillin
  • Growth Temperature
    30°C
  • Growth Strain(s)
    Stbl3
  • Growth instructions
    Use recombinase-free E. coli (Stbl3, XL-10, Sure2 et.al), grow at 30C
  • Copy number
    Low Copy

Gene/Insert

  • Gene/Insert name
    ArchT-tdTomato
  • Alt name
    archaerhodopsin
  • Alt name
    archaerhodopsin TP009
  • Alt name
    ArchT
  • Species
    H. strain TP009
  • Insert Size (bp)
    744
  • Mutation
    N/A
  • Promoter CAG
  • Tag / Fusion Protein
    • tdTomato (C terminal on insert)

Cloning Information

  • Cloning method Restriction Enzyme
  • 5′ cloning site KpnI (not destroyed)
  • 3′ cloning site BsrGI (not destroyed)
  • 5′ sequencing primer cttctggcgtgtgaccgg
  • (Common Sequencing Primers)

Resource Information

Depositor Comments

PLEASE CONTACT ED BOYDEN ([email protected]) FOR DETAILS ON THIS REAGENT AND FURTHER REAGENTS IN THIS LINE.

Information for AAV5 (Catalog # 28305-AAV5) ( Back to top )

Purpose

Ready-to-use AAV5 particles produced from pAAV-FLEX-ArchT-tdTomato (#28305). In addition to the viral particles, you will also receive purified pAAV-FLEX-ArchT-tdTomato plasmid DNA.

CAG-driven Cre-dependent ArchT-tdTomato expression for optogenetic neural inhibition. These AAV preparations are suitable purity for injection into animals.

Delivery

  • Volume 100 µL
  • Titer ≥ 7×10¹² vg/mL
  • Pricing $350 USD for preparation of 100 µL virus + $30 USD for plasmid.
  • Storage Store at -80℃. Thaw just before use and keep on ice.
  • Shipment Viral particles are shipped frozen on dry ice. Plasmid DNA (≥ 200ng) will also be included in the shipment.

Viral Production & Use

  • Packaging Plasmids encode adenoviral helper sequences and AAV rep gene, AAV5 cap gene
  • Buffer PBS + 0.001% Pluronic F-68
  • Serotype AAV5
  • Purification Iodixanol gradient ultracentrifugation
  • Reporter Gene tdTomato

Biosafety

Requestor is responsible for compliance with their institution's biosafety regulations. Lentivirus is generally considered BSL-2. AAV is generally considered BSL-1, but may require BSL-2 handling depending on the insert. Biosafety Guide

Resource Information

Viral Quality Control

Titering Method:
  • Real-time qPCR or digital droplet PCR: The number of genome copies in viral preparations was measured by real-time qPCR or by digital droplet PCR. Titering on these preparations was performed by the University of Pennsylvania Vector Core. Read Addgene's AAV Titration by qPCR protocol here.
Notes:
  • Purity of viral preparation: Viral preparations were subjected to polyacrylamide gel electrophoresis (PAGE) followed by SYPRO Red staining and the molecular weight and relative intensity of the viral capsid proteins was analyzed. The abundance of viral capsid proteins as a fraction of total protein present in the sample was used to determine purity of the AAV preparation.

Visit our viral production page for more information.

How to cite this plasmid ( Back to top)

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:

    pAAV-FLEX-ArchT-tdTomato was a gift from Edward Boyden (Addgene plasmid # 28305)

    For viral preps, please replace (Addgene plasmid # 28305) in the above sentence with: (Addgene viral prep # 28305-AAV5)

  • For your References section:

    A high-light sensitivity optical neural silencer: development and application to optogenetic control of non-human primate cortex. Han X, Chow BY, Zhou H, Klapoetke NC, Chuong A, Rajimehr R, Yang A, Baratta MV, Winkle J, Desimone R, Boyden ES. Front Syst Neurosci. 2011;5:18. Epub 2011 Apr 13. 10.3389/fnsys.2011.00018 PubMed 21811444