ABSTRACT
We developed a methylotrophic yeast, Pichia pastoris, as a host for DNA transformations. The system is based on an auxotrophic mutant host of P. pastoris which is defective in histidinol dehydrogenase. As a selectable marker, we isolated and characterized the P. pastoris HIS4 gene. Plasmid vectors which contained either the P. pastoris or the Saccharomyces cerevisiae HIS4 gene transformed the P. pastoris mutant host. DNA transfer was accomplished by a modified version of the spheroplast generation (CaCl2-polyethylene glycol)-fusion procedure developed for S. cerevisiae. In addition, we report the isolation and characterization of P. pastoris DNA fragments with autonomous replication sequence activity. Two fragments, PARS1 and PARS2, when present on plasmids increased transformation frequencies to 10(5)/micrograms and maintained the plasmids as autonomous elements in P. pastoris cells.
Subject(s)
Pichia/genetics , Saccharomycetales/genetics , Transformation, Genetic , Alcohol Oxidoreductases/genetics , Base Sequence , Chromosome Mapping , DNA, Fungal/genetics , Genes, Fungal , Mutation , Plasmids , Saccharomyces cerevisiae/geneticsABSTRACT
The cephalocarid crustacean Hutchinsoniella macracantha is a hermaphroditic species. Ova and sperm develop simultaneously. Ovaries and testes are separate, but the oviducts and vasa deferentia join and exit through a pair of common genital ducts.
Subject(s)
Crustacea/anatomy & histology , Genitalia/anatomy & histology , Animals , Disorders of Sex Development , Female , Male , Ovary , Oviducts , Testis , Vas DeferensABSTRACT
Normally acridine-sensitive, Escherichia coli-T2H complexes are rendered acridine-resistant if the infecting bacteriophage mutant is either pr or q. If these pr or q mutants are treated to produce sensitive revertants, one obtains a mutation at any of several dye-sensitizing (ds) sites in the early enzyme region of the T2 map. The ds mutants are nonspecific suppressors because they reduce the resistance of complexes containing either pr or q to proflavine. The ds mutants are not identical in action, since some make pr or q sensitive to proflavine and quinacrine, and others, to proflavine alone. Two ds mutants have r to r(+) mutation patterns which differ, depending upon whether or not the ds is coupled with r7 (an rII mutant). The mutation patterns of r(+) to r are the same for both ds mutants and for wild type. We suggest that dye sensitization may consist of alterations of early enzymes so as to produce slightly different forms of deoxyribonucleic acid which are in turn dyesensitive.
