Isolation and characterization of safflower (Carthamus tinctorius L.) chloroplast DNA

Safflower (Carthamus tinctorius L.) chloroplasts were isolated and purified with the aid of commercially available nylon mesh, differential centrifugation, and DNase I treatment. These chloroplasts were free of nuclei as determined by light microscopy of aceto orcein stained preparations, and similar to those observed by electron microscopy in spinach and many other higher plants, being bounded by a double membrane layer and characterized by the presence of a lamellar system surrounded by embedding matrix, and stacked membranes or grana lamallae. DNA was isolated and purified from such chloroplasts, and characterized with respect to cesium chloride density gradient isopycnic centrifugation, denaturation, renaturation kinetics and restriction enzyme analysis. These studies show that safflower chloroplast DNA is similar to many other higher plant DNAs having a density of 1.700 g/cm3 (G + C = 40.8%), a Tm of 86 degrees C (G + C = 40.7%) and a molecular complexity and genome size of about 10(8) daltons.