TRANSFECTION OF RABBIT ARTICULAR CHONDROCYTES BY THE CALCIUM-PHOSPHATE PROCEDURE - IMPROVEMENT OF EFFICIENCY AND REPRODUCIBILITY

The understanding of cartilage disorders relies on the possibility of studying mechanisms which monitor the regulation of matrix protein gen es through introducing efficiently and in a reproducible manner these genes, or their regulatory regions, into cells. To this end, we attemp ted to improve the transfection efficiency of rabbit articular chondro cytes by the calcium phosphate procedure. Transfection efficiencies we re assessed by measuring the expression of the Lac Z reporter gene enc oding P-galactosidase using an in situ staining (X-gal staining) and a n enzymatic assay (beta-galactosidase assay). Results revealed that ad dition of 4 U ml(-1) of hyaluronidase before and during transfection i ncreases by 2 to 4-fold the transfection efficiency of rabbit articula r chondrocytes. Furthermore, we demonstrated that the use of a ''giant '' calcium phosphate DNA coprecipitate gives a higher transfection eff iciency and much more reproducible results than those obtained with cl assical small volumes of precipitates.