The availability of gene constructs for various extracellular matrix molecules, makes it now possible to test the function of these molecules, the regulation of expression, and use in gene replacement therapy by incorporating these constructs into mouse eggs, thus creating transgenic mice. A transgenic mouse facility has been established and staff has been trained to carry out timed breeding of donor and foster mothers. The complex techniques involved in DNA injection have been mastered and injections are carried out on a daily routine. The collagen II constructs in which various lengths of the 5' flanking region are coupled to the gene for chloramphenicol acetyl transferase (CAT) with or without putative enhancer sequences have been used to produce a series of lines which differ in site of incorporation and copy number. Viable young which have incorporated the injected DNA into their chromosomes are then assayed for CAT activity to examine tissue specific expression. The BS-CAT construct (2Kb of the 5' flanking DNA from rat collagen II gene fused to CAT) has been successfully established as a stable transgenic line but has not shown the expected tissue and time specificity variations but in general demonstrate the expected activity in tissues that contain type II collagen. We anticipate that this system will be useful in testing recombinant constructs of cartilage and basement membrane genes as possible models for gene therapy. When full length cartilage proteoglycans are available, we plan to use mutant (CDM) mice unable to produce cartilage specific proteoglycan as a test system. Other mutant mouse lines such as congenital and juvenile polycystic kidney disease, hypochondroplasia, chondrodysplasia, progressive ankylosis, and fragilitas ossium are potential targets for this type of genetic intervention when the molecular basis of their defects is determined.
