Analysis of Euglena gracilis chloroplast deoxyribonucleic acid with a restriction endonuclease, EcoRI

Abstract

Cleavage of chloroplast DNA of E. gracilis Z with restriction endonuclease RI from Escherichia coli (EcoRI) yielded 23 bands on electrophoresis in gels of agarose. Four of the bands contained twice the stoichiometric amount of DNA. One of these bands contained 2 similarly sized fragments. The sum of the molecular weight of the 24 different fragments equaled the molecular weight of the circular molecule. The restriction fragments had different buoyant densities, with 4 having distinctly heavy densities in CsCl. Restriction fragments with a high buoyant density were preferentially lost when broken chloroplast DNA was purified by equilibrium density gradient centrifugation. Hybridization of chloroplast rRNA to intact chloroplast DNA determined that there are 2 cistrons for 16S and 23S rRNA. These 2 cistrons are located on 6 restriction fragments, all of which have buoyant densities greater than the intact molecule of chloroplast DNA.