Extended Data Fig. 5: The effect of DEN administration on the efficiency of gene targeting in mice liver.

a, DEN (10 or 30 mg/kg) was administered through a single i.p. injection per day for three days. Mice were also injected i.v. with rAAV8 packaged Alb-P2A-hF9 gene targeting vector (1.0 × 10¹¹ vg/mouse) on Day 1. Body weight was measured at the indicated time points. Data is displayed as the group mean with error bars representing s.d.; n = 4 mice per group. Significance testing was performed by two-way ANOVA with Dunnett’s multiple comparison test. P-value for DEN-10 treatment was .1589 and .0065 for DEN-30 treatment. b, Plasma hF9 protein levels in each treatment group was determined by ELISA. Data is displayed as the group mean with error bars representing s.d.; n = 4 mice per group. Significance testing was performed by one-way ANOVA with Dunnett’s multiple comparison test. P-value for DEN-10 treatment was .1301 and < .0001 for DEN-30 treatment. c-f, Total RNA was extracted from liver tissues and qPCR was performed to quantify the expression levels of (c) total hF9 mRNA, (d) endogenous albumin mRNA and (e) on-target integration derived Alb-P2A-hF9 fusion mRNA. (f) The fraction of hF9 fusion mRNA derived from on-target HR out of the total amount of hF9 mRNA is given. Actb mRNA was used for normalization and each data is shown as relative expression to the vehicle (saline)-injected control group. Data is displayed as the group mean with error bars representing s.d.; n = 4 mice per group. Significance was determined using two-tailed unpaired t tests for normally distributed data with equal variance or Welch’s correction for data with significantly different variance. Testing of non-normally distributed data was performed with a non-parametric Mann-Whitney U test.