FIG. 6.
Production of WISP3TG mice and demonstration of transgene expression in cartilage. (A) Schematic depicting the transgene construct containing WISP3 cDNA cloned with NotI restriction sites into the expression vector pKN185. PstI restriction sites and distances are noted. (B) Northern blot of costal cartilage (CC) and knee (KC) cartilage total RNA from WISP3TG and wild-type (WT) mice demonstrate detectable expression only in transgenic mice. (C) Southern blot of PstI-digested tail DNA probed with 32P-labeled full-length WISP3 cDNA showing 1-kb transgene fragments in a WISP3TG mouse but not in a WT mouse. WT mouse DNA spiked with transgene plasmid DNA prior to PstI digestion served as a positive control (+ control). (D) Western blot of protein extracts of femoral head cartilage from WISP3TG and WT mice after electrophoresis in a 10% SDS-Tris HCl gel under reducing conditions. Immunodetection with the polyclonal antibody (W3-C) demonstrates full-length (∼40 to 45 kDa) WISP3 protein in WISP3TG (arrowheads) but not WT mice.