Construction and mapping of safflower chloroplast DNA recombinants and location of selected gene markers

DNA was isolated and purified from chloroplasts of safflower (Carthamus tinctorius L.), digested with HindIII restriction endonuclease, and ligated into the HindIII site of the plasmid pUC9. Recombinant DNAs were isolated from ampicillin resistant white colonies which grew in the presence of the appropriate indicator, digested with HindIII, and then identified by comparison of agarose gel electrophoretic mobilities. HindIII digests of chloroplast DNA were used as a standard. Such recombinants were radiolabeled and hybridized with Southern blots of PstI, SalI, KpnI, and HindIII single and double digests of safflower chloroplast DNA. A physical map was subsequently generated showing the location of each recombinant on the circular plastid genome. Recombinants containing heterologous chloroplast gene markers from spinach or Euglena were also radiolabeled and mapped. The relative mapping positions of these genes are in good agreement with those which have previously been published for spinach and several other higher plants.