Cloning and sequencing of the ura3 and ura5 genes, and isolation and characterization of uracil auxotrophs of the fungus Mortierella alpina 1S-4.
The oil-producing fungus Mortierella alpina 1S-4 is an industrial strain. In order to prepare host strains for a transformation system for this fungus, six uracil auxotrophs were obtained by means of random mutation with N-methyl-N'-nitro-N-nitrosoguanidine (MNNG). When the activities of orotate phosphoribosyl transferase (OPRTase, EC 2.4.2.10) and orotidine-5'-phosphate decarboxylase (OMPdecase, EC 4.1.1.23) were examined in the mutants and wild strain, OPRTase activity was found to be completely absent in all mutants, on the other hand, OMPdecase activity was intact. The genomic DNA and cDNA of the ura5 gene encoding OPRTase and the ura3 gene encoding OMPdecase were cloned and sequenced. The Ura5p deduced amino acid sequence of this fungus showed highest similarity to that of Vibrio cholerae classed among prokaryote. Furthermore, the mutational points in the ura5 genes of two selected mutants were identified; a base-replacement and a base-insertion.
Laboratory of Fermentation Physiology and Applied Microbiology, Division of Applied Life Sciences, Graduate School of Agriculture, Kyoto University, Japan.
Seiki
S
Takeno
Eiji
E
Sakuradani
Shoichi
S
Murata
Misa
M
Inohara-Ochiai
Hiroshi
H
Kawashima
Toshihiko
T
Ashikari
Sakayu
S
Shimizu
D000595:Amino Acid Sequence; D001483:Base Sequence; D015139:Blotting, Southern; D003001:Cloning, Molecular; D003470:Culture Media; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D004271:DNA, Fungal / Q000096:biosynthesis / Q000235:genetics; D015723:Gene Library; D005800:Genes, Fungal / Q000235:genetics*; D008969:Molecular Sequence Data; D020094:Mortierella / Q000235:genetics* / Q000378:metabolism; D009154:Mutation / Q000235:genetics; D009821:Oils / Q000378:metabolism; D009962:Orotate Phosphoribosyltransferase / Q000235:genetics*; D009964:Orotidine-5'-Phosphate Decarboxylase / Q000235:genetics*; D014498:Uracil / Q000737:chemistry / Q000378:metabolism*
D003470:Culture Media; D018076:DNA, Complementary; D004271:DNA, Fungal; D009821:Oils; D014498:Uracil; D009962:Orotate Phosphoribosyltransferase; D009964:Orotidine-5'-Phosphate Decarboxylase