title: Identification and application of novel autonomously replicating sequences (ARSs) for promoter-cloning and co-transformation in Candida utilis.
abstract: In order to develop practical recombinant DNA techniques in the industrially important yeast Candida utilis, at least six plasmids harboring autonomously replicating sequences (ARSs) were isolated from a C. utilis genomic library. Two ARSs were subjected to detailed analysis. Sequences of 1.9 and 1.8 kb were found to be necessary to exert ARS activity in a plasmid as assessed by transformation efficiency and mitotic stability. Both fragments were found to be rich in AT content (69.5% and 70.8% respectively), and to contain an 11-bp ARS consensus sequences (10 and 13 motifs with one base difference respectively). Using the ARS-containing plasmid as a promoter-cloning vector, several DNA fragments having promoter activities were cloned and characterized. Co-transformation of C. utilis with an integrating DNA fragment and a replicating plasmid yielded plasmid-free transformants harboring the fragment integrated into the C. utilis genome.
authors.affiliation: Central Laboratories for Frontier Technology, KIRIN Holdings Co., Ltd. Shigehito_Ikushima@kirin.co.jp
authors.forename: Shigehito
authors.initials: S
authors.lastname: Ikushima
authors.forename: Toshiko
authors.initials: T
authors.lastname: Minato
authors.forename: Keiji
authors.initials: K
authors.lastname: Kondo
mesh_terms: D002175:Candida / Q000235:genetics*; D003001:Cloning, Molecular; D016384:Consensus Sequence / Q000235:genetics; D004261:DNA Replication; D004274:DNA, Recombinant; D016681:Genome, Fungal / Q000235:genetics; D010957:Plasmids; D011401:Promoter Regions, Genetic; D014170:Transformation, Genetic
chemical_list: D004274:DNA, Recombinant