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Validation Experiment: Independent validation of Deverman delivery platform using engineered AAVs to deliver CRSIPR/Cas9 to mouse brain

PI:   Jason D Heaney, PhD 

Description: Validation of delivery of AAV custom designed to cross the blood-brain barrier for CRISPR/Cas9 editing. Editing detected and quantified in brain by generation of tdTomato fluorescent protein signal from Ai9 reporter mice
Delivery Assays: Identification of of GFP-positive nuclei (green)
Editing Assay:
  • Qualitative assesment of tdTomato-negative cells throughout the entire brain
  • Quantitative assessment of tdTomato-positive cells per DAPI-stained nucleus in a specific brain region
  • Qualitative assesment of prevalence of tdTomato-positvie cells throughout the entire brain
Download: Submitted files
Original Experiment/s that are being validated:
Filter Results ..
  Tissues
Nervous
  • Brain (neuron)  (TARGET)
  • Cerebral cortex (astrocyte)  (TARGET)
  • Dorsal plus ventral thalamus (astrocyte)  (TARGET)
  • Brain (astrocyte)  (TARGET)
  • Brain  (TARGET)
  Cell Types
 astrocyte
 neuron
Editors
 SaCas9
Guide Target Locus
 Ai9/Ai14 reporter transgene
  Guides
 L1-modified
 L2-modified
Models
 Ai9 mouse
  Sex
 Female
 Male
  Vectors
 BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2
 BI28:AAV-GFAP-SaCas9-WPRE3-pA
Select experimental variable to highlight records on the chart:

Note: Hover over the bars to view additional information

Results
Independent_validation_of_Deverman_delivery_platform_using_engineered_AAVs_to_deliver_CRSIPR/Cas9_to_mouse_brain
Editing Efficiency Delivery Efficiency
Record Id Condition Tissue Cell Type Sex Editor Model Target Locus Guide Vector Dosage Injection Frequency Editing Efficiency Signal Editing Efficiency % Area Delivery Efficiency Signal Image
15000001448 Arm1a: BI28 Neuron Specific Promoter, Experimental Animals, Males brain (TARGET) neuron Male SaCas9 Ai9 mouse Ai9/Ai14 reporter transgene L1-modified; L2-modified BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2; BI28:AAV-GFAP-SaCas9-WPRE3-pA 0.25E+11 viral genomes/gram for each virus once
15000001449 Arm1a: BI28 Neuron Specific Promoter, Experimental Animals, Males brain (TARGET) neuron Male SaCas9 Ai9 mouse Ai9/Ai14 reporter transgene L1-modified; L2-modified BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2; BI28:AAV-GFAP-SaCas9-WPRE3-pA 0.25E+11 viral genomes/gram for each virus once
15000001450 Arm1a: BI28 Neuron Specific Promoter, Experimental Animals, Females brain (TARGET) neuron Female SaCas9 Ai9 mouse Ai9/Ai14 reporter transgene L1-modified; L2-modified BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2; BI28:AAV-GFAP-SaCas9-WPRE3-pA 0.25E+11 viral genomes/gram for each virus once
15000001451 Arm1a: BI28 Neuron Specific Promoter, Experimental Animals, Females brain (TARGET) neuron Female SaCas9 Ai9 mouse Ai9/Ai14 reporter transgene L1-modified; L2-modified BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2; BI28:AAV-GFAP-SaCas9-WPRE3-pA 0.25E+11 viral genomes/gram for each virus once
15000001452 Arm 1b: BI28 Neuron Specific Promoter, Positive Delivery Control Animals, Males brain (TARGET) neuron Male SaCas9 Ai9 mouse BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2 0.25E+11 viral genomes/gram for each virus once
15000001453 Arm 1b: BI28 Neuron Specific Promoter, Positive Delivery Control Animals, Females brain (TARGET) neuron Female SaCas9 Ai9 mouse BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2 0.25E+11 viral genomes/gram for each virus once
15000001454 Arm 2a: BI28 Astrocyte Specific Promoter, Experimental Animals, Males cerebral cortex (TARGET) astrocyte Male SaCas9 Ai9 mouse Ai9/Ai14 reporter transgene L1-modified; L2-modified BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2; BI28:AAV-GFAP-SaCas9-WPRE3-pA 0.25E+11 viral genomes/gram for each virus once
15000001455 Arm 2a: BI28 Astrocyte Specific Promoter, Experimental Animals, Males dorsal plus ventral thalamus (TARGET) astrocyte Male SaCas9 Ai9 mouse Ai9/Ai14 reporter transgene L1-modified; L2-modified BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2; BI28:AAV-GFAP-SaCas9-WPRE3-pA 0.25E+11 viral genomes/gram for each virus once
15000001456 Arm 2a: BI28 Astrocyte Specific Promoter, Experimental Animals, Males brain (TARGET) astrocyte Male SaCas9 Ai9 mouse Ai9/Ai14 reporter transgene L1-modified; L2-modified BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2; BI28:AAV-GFAP-SaCas9-WPRE3-pA 0.25E+11 viral genomes/gram for each virus once
15000001457 Arm 2a: BI28 Astrocyte Specific Promoter, Experimental Animals, Females cerebral cortex (TARGET) astrocyte Female SaCas9 Ai9 mouse Ai9/Ai14 reporter transgene L1-modified; L2-modified BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2; BI28:AAV-GFAP-SaCas9-WPRE3-pA 0.25E+11 viral genomes/gram for each virus once
15000001458 Arm 2a: BI28 Astrocyte Specific Promoter, Experimental Animals, Females dorsal plus ventral thalamus (TARGET) astrocyte Female SaCas9 Ai9 mouse Ai9/Ai14 reporter transgene L1-modified; L2-modified BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2; BI28:AAV-GFAP-SaCas9-WPRE3-pA 0.25E+11 viral genomes/gram for each virus once
15000001459 Arm 2a: BI28 Astrocyte Specific Promoter, Experimental Animals, Females brain (TARGET) astrocyte Female SaCas9 Ai9 mouse Ai9/Ai14 reporter transgene L1-modified; L2-modified BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2; BI28:AAV-GFAP-SaCas9-WPRE3-pA 0.25E+11 viral genomes/gram for each virus once
15000001460 Arm 2b: BI28 Astrocyte Specific Promoter, Positive Delivery Control Animals, Males brain (TARGET) astrocyte Male SaCas9 Ai9 mouse BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2 0.25E+11 viral genomes/gram for each virus once
15000001461 Arm 2b: BI28 Astrocyte Specific Promoter, Positive Delivery Control Animals, Females brain (TARGET) astrocyte Female SaCas9 Ai9 mouse BI28:AAV-GFAP-NLS-GFP-WPRE-synpA-L1-R2 0.25E+11 viral genomes/gram for each virus once
15000001462 Arm 3: Negative Control Saline, Males brain (TARGET) Male Ai9 mouse once
15000001463 Arm 3: Negative Control Saline, Males brain (TARGET) Male Ai9 mouse once

Experiment Wide Protocols

Title Description File Download SCGE ID
Heaney_SATC Tissue Processing, Imaging and Analysis Procedure for tissue preparation, imaging and analysis. Heaney_SATC_Tissue_Processing_Imaging_and_Analysis.pdf 21000000025