Amplification of Hot DNA segments in Escherichia coli
Amplification of Hot DNA segments in Escherichia coli
复制标题
大肠杆菌中热 DNA 片段的扩增
DOI：
10.1046/j.1365-2958.2002.03141.x
复制
发表时间：
2002
影响因子：
3.6
通讯作者：
T. Horiuchi
中科院分区：
文献类型：
--
作者：
K. Kodama;Takehiko Kobayashi;H. Niki;S. Hiraga;T. Oshima;H. Mori;T. Horiuchi
In Escherichia coli, a replication fork blocking event at a DNA replication terminus (Ter) enhances homologous recombination at the nearby sister chromosomal region, converting the region into a recombination hotspot, Hot, site. Using a RNaseH negative (rnhA–) mutant, we identified eight kinds of Hot DNAs (HotA–H). Among these, enhanced recombination of three kinds of Hot DNAs (HotA–C) was dependent on fork blocking events at Ter sites. In the present study, we examined whether HotA DNAs are amplified when circular DNA (HotA plus a drug‐resistance DNA) is inserted into the homologous region on the chromosome of a rnhA– mutant. The resulting HotA DNA transformants were analysed using pulsed‐field gel electrophoresis, fluorescence in situ hybridization and DNA microarray technique. The following results were obtained: (i) HotA DNA is amplified by about 40‐fold on average; (ii) whereas 90% of the cells contain about 6–10 copies of HotA DNA, the remaining 10% of cells have as many as several hundred HotA copies; and (iii) amplification is detected in all other Hot DNAs, among which HotB and HotG DNAs are amplified to the same level as HotA. Furthermore, HotL DNA, which is activated by blocking the clockwise oriC‐starting replication fork at the artificially inserted TerL site in the fork‐blocked strain with a rnhA+ background, is also amplified, but is not amplified in the non‐blocked strain. From these data, we propose a model that can explain production of three distinct forms of Hot DNA molecules by the following three recombination pathways: (i) unequal intersister recombination; (ii) intrasister recombination, followed by rolling‐circle replication; and (iii) intrasister recombination, producing circular DNA molecules.
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DOI：
--
发表时间：
2000
期刊：
Genes & development.
影响因子：
--
作者：
Rothstein,R;Michel,B;Gangloff,S
通讯作者：
Gangloff,S
大 DNA 分子的纯化、特异性片段化和分离。
影响因子：
--
作者：
Smith,CL;Cantor,CR
通讯作者：
Cantor,CR
影响因子：
56.9
作者：
Andersson, DI;Slechta, ES;Roth, JR
通讯作者：
Roth, JR