RATE, ORIGIN, AND BIDIRECTIONALITY OF CAULOBACTER CHROMOSOME-REPLICATION AS DETERMINED BY PULSED-FIELD GEL-ELECTROPHORESIS
RATE, ORIGIN, AND BIDIRECTIONALITY OF CAULOBACTER CHROMOSOME-REPLICATION AS DETERMINED BY PULSED-FIELD GEL-ELECTROPHORESIS
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DOI：
10.1073/pnas.86.1.119
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发表时间：
1989-01-01
影响因子：
11.1
通讯作者：
SHAPIRO, L
中科院分区：
文献类型：
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作者：
DINGWALL, A;SHAPIRO, L
新月柄杆菌的细胞分裂产生的子代细胞在细胞结构和发育程序方面不同。染色体复制起始于子柄细胞，但在子群细胞中受到抑制，直到细胞周期的后期。为了研究细胞类型特异性DNA起始，通过脉冲场凝胶电泳直接分析染色体复制。在不同时间从用2“-脱氧[3 H]-鸟苷标记的同步化细胞培养物中提取DNA的Dra I限制性片段，分析这些片段使我们能够确定DNA复制的起点、分叉运动的速率和方向以及基因复制的顺序。出现的第一个标记的Dra I片段含有复制起始位点。根据伊利和Gerardot [伊利，B.和Gerardot，C. J.（1988）Gene 68，323-333]，起源定位于含有rrnA基因的305-内切酶片段。此外，通过未映射的Dra I片段的连续复制使我们能够定位它们在基因组上的位置。标记的限制性片段的出现顺序表明，染色体以每分钟21个酶的叉移动速率双向复制。
Cell division in Caulobacter crescentus yields progeny cells that differ with respect to cell structure and developmental program. Chromosome replication initiates in the daughter stalked cell but is repressed in the daughter swarmer cell until later in the cell cycle. To study cell-type-specific DNA initiation, chromosome replication was directly analyzed by pulsed-field gel electrophoresis. Analysis of Dra I restriction fragments of DNA taken at various times from synchronized cell cultures labeled with 2''-deoxy[3H]-guanosine has allowed us to determine the origin of DNA replication, the rate and direction of fork movement, and the order of gene replication. The first labeled Dra I fragment to appear contains the site of replication initiation. Based on the correlation of the physical and genetic maps derived by Ely and Gerardot [Ely, B. and Gerardot, C. J. (1988) Gene 68, 323-333], the origin was localized to a 305-kilobase fragment containing the rrnA gene. Furthermore, the sequential replication through unmapped Dra I fragments has enabled us to localize their positions on the genome. The order of appearance of labeled restriction fragments revealed that the chromosome replicates bidirectionally at a fork movement rate of 21 kilobases per minute.