Figure 1
Generation of human IL6 receptor (IL6R) gene knock-in mouse.
(a) Schematic representation of the knock-in strategy for the hIL6R gene. A knock-in vector was constructed by inserting hIL6R cDNA with neo cassette flanked by two loxP sites into the mouse Il6ra genomic locus in the frame of a BAC genomic clone. A knock-in allele and a neo-deleted knock-in allele are also shown. Arrows indicate PCR primers (m10355 and m11166R) for genotyping. TC, terminal codon. (b) A representative result of genotyping to confirm the neo-depleted hIL6R knock-in allele and homozygosity of the hIL6R knock-in allele. Wild-type allele and knock-in allele were detected as signals of 0.8 kb and 4.2 kb, respectively, whereas knock-in allele after removing neo cassette was detected as a signal of 2.7 kb. M, DNA molecular marker. Numbers above the gel denote the mouse genotypes, (1) Il6ra+/+, (2) Il6rahIL6R/+, (3) Il6rahIL6R/hIL6R and (4) Il6rahIL6R(neo)/hILR6(neo). (c) Representative results of RT-PCR analysis for tissue distribution of Il6ra+/+ (Wi) and Il6rahIL6R/hIL6R (Ho) mice. (d) Plasma levels of soluble hIL6R in Il6ra+/+ (n = 14), Il6rahIL6R/+ (n = 16) and Il6rahIL6R/hIL6R mice (n = 13). (e) Species-specific ligand response was confirmed after intraperitoneal injection of mouse Il6 (mIL6) or human IL6 (hIL6) in Il6ra+/+ and in Il6rahIL6R/hIL6R mice. Ligand responses were evaluated by the elevation of plasma SAA levels after injection of vehicle (n = 3), mIL6 (n = 4) and hIL6 (n = 3) in Il6ra+/+ and those of vehicle (n = 2), mIL6 (n = 3) and hIL6 (n = 3) in Il6rahIL6R/hIL6R.
