A DNA-mediated transformation system for the blue-green alga Agmenellum quadruplicatum, strain PR-6, is described and characterized for DNA concentration dependence, dependence on time of exposure to DNA, phenotypic expression, sensitivity to various enzymes, and competence. The stability of the transformants has been investigated, and genetic backcross and selfing experiments have been performed. This system fulfills all of the criteria established for the well-characterized transformation systems in heterotrophic bacteria and demonstrates significant similarities to at least one of these systems for all characteristics examined. The efficiency of transformation is high. This system fills a need for a well-characterized genetic system in an oxygen-evolving photoautotroph. We have used it to transform a strain with a mutational lesion in assimilatory nitrogen metabolism to a wild-type genotype. The blue-green algae (cyanobacteria) provide an excellent model system for the study of photoautotrophic metabolism. They carry out oxygen-evolving photosynthesis and assimilate oxidized nitrogen in a fashion similar to that of higher plants and are amenable to many of the techniques utilized in the study of heterotrophic bacteria. A limiting factor in studying the metabolism of these organisms has been the lack of a system for genetic exchange. Although there have been reports of genetic exchange in blue-green algae (reviewed in ref. 1), only the transformation system in Anacystis nidulans has been reasonably well characterized (2, 3).We have found that the blue-green alga Agmenellum quadruplicatum, strain PR-6, possesses an efficient, naturally occurring mechanism for the uptake and integration of exogenous DNA in a process like transformation in other bacteria (4). Here we characterize this system for sensitivity to DNase, dependence on concentration of DNA, dependence on time of exposure of cells to DNA, competence, and expression of newly incorporated genetic material. The PR-6 transformation system is similar to the standard heterotrophic bacterial transformation systems in all respects thus far tested.We also describe an initial experiment wherein a physiologically well-characterized mutant of PR-6, called AQ-6 (5-8), was transformed with parental DNA. AQ-6 reduces nitrate to nitrite but its reduction of nitrite is impaired, resulting in the accumulation of nitrite in the medium. The mutant grows normally in the presence of ammonia. After transformation at least two colonial phenotypes were recovered. Genetic dissection of inorganic nitrogen assimilation in blue-green algae is now feasible. Growth and cell concentration of PR-6 and AQ-6 were routinely measured turbidimetrically with a Spectronic 20 colorimeter at 550 nm. An average concentration of 4.2 X 107 cells per ml was indicated by an optical density of 0.82 in a 22-mm culture tube. MATERIALS AND METHODSDNA Preparation. DNA from and from a streptomycin-resistant derivative (see below) were purified by the procedure of Marmur (11).Standard Transforma...