Figure S5.
The validation of AAV9 gene delivery in the primary motor cortex region of Arl6ip1−/−and Arl6ip1+/+mice. (A) Schematic representation of the AAV9-CAG-GFP-ARL6IP1 plasmid and the BamHI restriction sites. Representative image of gel electrophoresis of an undigested and digested plasmid. (B) After BamHI digestion, the purity and concentration of AAV9-CAG-GFP-ARL6IP1 plasmid were measured using a nanodrop spectrophotometer. (C) AAV9-CAG-GFP-ARL6IP1 plasmid was serially diluted and analyzed by qPCR using EGFP and SV40 primers. A qPCR standard curve was created by plotting Ct values against the corresponding Log DNA copies/ml. (D and E) Representative immunofluorescence images and quantification of GFP, and GFAP in the primary motor cortex region of Arl6ip1−/− mice after AAV9 gene delivery (200×; scale bars: 100 μm). (F and G) Representative immunofluorescence images and quantification of GFP, and GFAP in the primary motor cortex area of Arl6ip1+/+ mice after AAV9 gene delivery (200×; scale bars: 100 μm). Data (E and G) represent mean ± SD of N = 5 mice/group and the two-tailed unpaired t test. *P < 0.05, **P < 0.01. (H) After AAV9 gene delivery, Arl6ip1, and EGFP mRNA levels were confirmed for gene transfer efficiency in the cerebral cortex of Arl6ip1+/+ mice. mRNA levels of glial or pro-inflammatory marker genes after AAV9 gene delivery. Data are represented with the average of triple technical repeats, the mean and error bars represent the SEM, and the two-tailed unpaired t test. *P < 0.01, **P < 0.001 (n = 3 mice/group). All experiments were performed in three independent biological replicates. Source data are available for this figure: SourceData FS5.

The validation of AAV9 gene delivery in the primary motor cortex region of Arl6ip1 −/− and Arl6ip1 +/+ mice. (A) Schematic representation of the AAV9-CAG-GFP-ARL6IP1 plasmid and the BamHI restriction sites. Representative image of gel electrophoresis of an undigested and digested plasmid. (B) After BamHI digestion, the purity and concentration of AAV9-CAG-GFP-ARL6IP1 plasmid were measured using a nanodrop spectrophotometer. (C) AAV9-CAG-GFP-ARL6IP1 plasmid was serially diluted and analyzed by qPCR using EGFP and SV40 primers. A qPCR standard curve was created by plotting Ct values against the corresponding Log DNA copies/ml. (D and E) Representative immunofluorescence images and quantification of GFP, and GFAP in the primary motor cortex region of Arl6ip1−/− mice after AAV9 gene delivery (200×; scale bars: 100 μm). (F and G) Representative immunofluorescence images and quantification of GFP, and GFAP in the primary motor cortex area of Arl6ip1+/+ mice after AAV9 gene delivery (200×; scale bars: 100 μm). Data (E and G) represent mean ± SD of N = 5 mice/group and the two-tailed unpaired t test. *P < 0.05, **P < 0.01. (H) After AAV9 gene delivery, Arl6ip1, and EGFP mRNA levels were confirmed for gene transfer efficiency in the cerebral cortex of Arl6ip1+/+ mice. mRNA levels of glial or pro-inflammatory marker genes after AAV9 gene delivery. Data are represented with the average of triple technical repeats, the mean and error bars represent the SEM, and the two-tailed unpaired t test. *P < 0.01, **P < 0.001 (n = 3 mice/group). All experiments were performed in three independent biological replicates. Source data are available for this figure: SourceData FS5.

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