The codon-optimized human FIX cDNA (co-hFIX) or its hyper-functional codon-optimized version (co-hFIX-R338L) were cloned downstream of a chimeric liver-specific promoter composed on the minimal transthyretin (TTR) promoter in combination with a hepatocyte-specific cis-regulatory module

Vector constructs The codon-optimized human FIX cDNA (co-hFIX) or its hyper-functional codon-optimized version (co-hFIX-R338L) were cloned downstream of a chimeric liver-specific promoter composed on the minimal transthyretin (TTR) promoter in combination with a hepatocyte-specific cis-regulatory module (designated as HS-CRM8) that was designed in silico, as described 1,2. This HS-CRM8 contains evolutionary conserved clusters of transcription factor binding sites (TFBS) associated with high liver-specific gene expression. This expression cassette was cloned into a self-complementary (sc)AAV backbone 3. The vectors also contained a mini-intron from minute virus of mice (MVM) and a bovine growth hormone polyadenylation site (bGH polyA) (Figure 1). To generate the AAV-HS-CRM8-TTR-GFP vector the green fluorescent protein (GFP) gene was cloned downstream from HS-CRM8/TTR. Cloning details are available upon request.