• Parkinsons Disease AAV Research Tools


    MJFF Image

    Vigene has partnered with The Michael J. Fox Foundation for Parkinson’s Research (MJFF) to distribute viral vectors targeting alpha-synuclein, a major protein implicated in the pathology of Parkinson’s disease (PD).
    Learn more about alpha-synuclein and PD.

    alpha-synuclein AAV viral vectors

    aSyn knockdown: Viral vectors expressing a miR to specifically knockdown mouse aSyn, human aSyn, or a scrambled control, as well as a GFP reporter to measure transduction efficiency.

    aSyn overexpression: Two different AAV pseudotypes to overexpress wildtype human aSyn—rAAV2/2 aSyn and rAAV2/5 aSyn—with rAAV2/2 BFP and rAAV2/5 BFP as their respective controls.

    alpha-synuclein knockdown viral vectors: mouse and human

    In collaboration with MJFFs Parkinson's Disease Research Tools Consortium, three AAV viral vectors were generated that knockdown the levels of aSyn for use as a preclinical tool to understand alpha-synuclein biology. These viral vectors each express a miR to specifically knockdown mouse aSyn, human aSyn, or a scrambled control, as well as a GFP reporter to measure transduction efficiency. With the help of MJFF industry partners, these vectors are fully validated for expression and knockdown efficiency in vitro as well as in vivo. View validation data.

    Knockdown efficiency: in vitro ~75%; in vivo ~100%

    Volume: 100µl each

    Titer: 0.5x1012 vg/mL

    Price: $250

    Diluent: rAVE-Vector Reagent download datasheet

    Order alpha-synuclein AAV tools

    Cat #

    aSyn

    AAV Vector

    Download

    Order

    GD1009-RV-H

    human

    AAV1/2-CAG-Human-SNCA-3xmiR/GFP-WPRE-BGH-polyA

    datasheet or
    NTI sequence

    Order

    GD1009-RV-M

    mouse

    AAV1/2-CAG-Mouse-SNCA-3xmiR/GFP-WPRE-BGH-polyA

    datasheet or
    NTI sequence

    Order

    GD1009-RV-C

    scrambled control

    AAV1/2-CAG-Scrambled-Control-3xSCRmiR/GFP-WPRE-BGH-polyA

    datasheet or
    NTI sequence

    Order

    Recommended Dose

    For in vitro use GeneDetect recommend starting using AAV at 1 in 100 dilution of concentrate AAV solution supplied at 0.5 x 10^12 vg/ml. Dilute further as necessary. Wait 2-3 days for HEK293 cells, 7-9 days for primary neurons.

    For in vivo use GeneDetect recommend starting with a 2µl injection of concentrate AAV solution supplied at 0.5 x 10^12 vg/ml. Dilute further as necessary. Wait three weeks, generally.

    All cell types are different but 1:100 is a good starting dose and 2µl of full strength AAV into brain obliterates alpha synuclein expression.

    Validation data

    Knockdown efficiency of the constructs in transiently transfected HEK293 cells

    Transduction efficiency figure

    A-B aSyn immunoreactivity in HEK cells 24hrs after co-transfection with human or mouse aSyn-expressing constructs and empty constructs, GFP constructs, or SNCA miR-GFP constructs. Scale bars = 250μm. A HEK293 cells successfully overexpress human or mouse aSyn, with knockdown of this expression resulting from co-transfection with the associated SNCA miR. B Transfection with an empty plasmid instead of an aSyn-expressing plasmid does not result in aSyn expression. C-D Western blot detection and quantitation of aSyn and GFP in lysates from the HEK293 cells in panels A and B. The mouse SNCA miR construct significantly reduces expression of mouse aSyn without affecting human aSyn expression. The human SNCA miR construct significantly reduces human aSyn expression with some cross-reactivity resulting in a reduction of mouse aSyn. The GFP construct reduced human aSyn expression but not mouse aSyn expression, indicating that high levels of GFP expression may attenuate the low levels of human aSyn expression. GAPDH was the loading control. Bars represent mean ± SEM (n=3 per treatment). * p < 0.05, *** p < 0.001, **** p < 0.0001 by one-way ANOVA with Tukey’s post-hoc analysis. Viral vectors were designed, generated, and validated by GeneDetect.

    Transduction efficiency and aSyn knockdown

    The figure below shows the transduction efficiency and aSyn knockdown after co-infusion of aSyn-overexpressing viral vectors with SNCA miR/GFP viral vectors in rat substantia nigra pars compacta (SNC).

    Transduction efficiency figure

    A-B Validation of transduction efficiency with the human aSyn-overexpressing viral vector and the scrambled control miR viral vector at three weeks post-co-infusion. Scale bar = 500µm, arrow indicates aSyn overexpression in the SNC. A aSyn-overexpressing viral vectors result in detectable increases in aSyn protein in the SNC three weeks post-injection. B The scrambled control miR/GFP viral vector robustly expresses the transgene in the SNC. C-D Co-infusion of the human or mouse aSyn-overexpressing viral vector with the various miR viral vectors. Scale bars = 100μm, arrows indicate aSyn-positive cells in the SNC adjacent to the substantia nigra pars reticulata (SNR).
    C Human aSyn overexpression was abolished by the human SNCA miR/GFP viral vector, slightly reduced with the mouse SNCA miR/GFP viral vector, and unaltered with the scrambled control miR/GFP viral vector. D Mouse aSyn overexpression was abolished by the mouse SNCA miR/GFP viral vector as well as the human SNCA miR/GFP viral vector, with no appreciable decreases in mouse aSyn overexpression observed with the scrambled control miR/GFP. Viral vectors were designed, generated, and validated by GeneDetect.

    Alpha-synuclein overexpression viral vectors

    Ready to ship mid-late 2019

    Alpha–synuclein

    Alpha-synuclein (a-syn) is a protein whose function in the healthy brain is currently unknown. Alpha-synuclein is of great interest to Parkinson's researchers because it is a major constituent of Lewy bodies, or protein clumps that are the pathological hallmark of Parkinson's disease. There is compelling evidence from recent studies that aSyn may play a role in the development of both familial (rare) and sporadic (more common) cases of PD.

    Alpha–synuclein Therapy

    A vaccine approach targeting alpha-synuclein in people with Parkinson's is now in the clinic. The vaccine candidate, from Austrian biotech AFFiRiS, works by binding to alpha-synuclein and subsequently clearing it from the brain. It is now in the first stages of clinical testing. Other approved compounds that have been shown to stop synuclein clumping and break up existing clumps are moving forward in pre-clinical studies.

    The Michael J. Fox Foundation Pre-clinical Tools Program

    To speed Parkinson’s research and accelerate therapeutic development, MJFF invests in the generation and availability of critical tools for Parkinson’s disease investigators. MJFF has a multi-faceted approach to developing these research tools. We help identify critical needs through discussions with key scientific experts, generate tools through partnerships with leading experts and companies, evaluate these resources for quality control, and distribute the tools broadly through vendors and partner organizations. In addition, the Foundation is spearheading the Parkinson's Disease Research Tools Consortium, working with industry partners to develop and characterize new tools to address unmet challenges in PD research. More information on MJFF tools and partnerships and can be found in the MJFF Research Tools Catalog.

    Additional Viral Resources

    Ordering Instructions: Shipping and payment information
    Product Manuals: How to use your virus
    FAQs : Answers to commonly asked questions
    Publications: showcasing Vigene products and services
    Recipient instructions: information on viral safety and storage