Purification of plastid DNA from an enriched rhodoplast fraction of the red alga Gracilaria tenuistipitata

We report a straightforward protocol for isolating plastid DNA from an enriched rhodoplast fraction of the red alga Gracilaria tenuistipitata. Plastids were purified using differential centrifugation and 2-step sucrose density gradients. We found that 10% polyethylene glycol 4000 was essential for maintaining plastid integrity prior to lysis. Plastid DNA isolated directly from the purified rbodoplasts was sufficiently pure for restriction endonuclease fragment analyses. Database comparisons of sequences generated randomly from a shotgun genomic library indicated that plastid DNA was 89% pure following ultracentrifugation in isopycnic cesium chloride equilibrium gradients. The protocol yields 30-50 mug of plastid DNA per 100 g of fresh algal tissue.