Figure 3.

Indel and HDR-Mediated Gene Integration Efficiency and Transgene Expression Analyses

Liver mRNA and DNA were isolated from hemophilia B mice at 14, 16, and 48 weeks after treatment with dual gene integration vectors or untargeted vectors. DNA and mRNA from untreated hemophilia B mice served as control. Indel analysis on the mAlb locus was analyzed by NGS on liver DNA. HDR-mediated gene integration was analyzed by LM-PCR following digestion with XbaI on liver DNA. The pooled PCR amplicons were performed by NGS. Chimeric m-hFIX mRNA levels in liver were measured by qPCR using primers spanning the junction of mAlb and hFIX cDNA. (A) Indel frequency in mice treated as adults. (B) Indel frequency in mice treated as neonates. (C) HDR frequency in mice treated as adults. (D) HDR frequency in mice treated as neonates. (E) Quantification of chimeric m-hFIX mRNA levels in the liver treated as adults and neonates. Symbols represent individual mice. Means ± SEM are shown. Dunnett’s test. n.s., not significant.