Leiting B, Noegel A
Max-Planck-Institut für Biochemie, Martinsried, Federal Republic of Germany.
Plasmid. 1988 Nov;20(3):241-8. doi: 10.1016/0147-619x(88)90030-3.
An extrachromosomally replicating transformation vector for Dictyostelium discoideum has been constructed using sequences of the endogenous Dictyostelium plasmid Ddp2. This transformation vector pnDeI (9.6 kb) replicates as a high copy number plasmid in Dictyostelium and is located in the nucleus. It has been constructed as shuttle vector containing the Escherichia coli vector pUC19 for replication and selection in E. coli and a part of the Tn903 transposon which confers resistance to G418 for selection in Dictyostelium. In order to show that the vector can be used for cloning and stable propagation of Dictyostelium DNA, a fragment of the Dictyostelium alpha-actinin gene that was marked with a synthetic oligonucleotide was cloned into pnDeI and found to be stably maintained in the extrachromosomal vector without undergoing noticeable recombination with the endogenous gene.
利用盘基网柄菌内源性质粒Ddp2的序列构建了一种用于盘基网柄菌的染色体外复制转化载体。这种转化载体pnDeI（9.6 kb）在盘基网柄菌中作为高拷贝数质粒进行复制，并且定位于细胞核中。它被构建为穿梭载体，包含用于在大肠杆菌中复制和筛选的大肠杆菌载体pUC19，以及赋予对G418抗性以便在盘基网柄菌中筛选的Tn903转座子的一部分。为了证明该载体可用于盘基网柄菌DNA的克隆和稳定增殖，将用合成寡核苷酸标记的盘基网柄菌α-肌动蛋白基因片段克隆到pnDeI中，发现其在染色体外载体中稳定维持，而未与内源性基因发生明显重组。
