The dynamics of genome replication using deep sequencing.
The dynamics of genome replication using deep sequencing.
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DOI：
10.1093/nar/gkt878
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发表时间：
2014-01
影响因子：
14.9
通讯作者：
Nieduszynski CA
中科院分区：
文献类型：
--
作者：
Müller CA;Hawkins M;Retkute R;Malla S;Wilson R;Blythe MJ;Nakato R;Komata M;Shirahige K;de Moura AP;Nieduszynski CA
真核生物基因组从多个DNA复制起点复制。我们提出了互补的深度测序方法来测量酿酒酵母中的起源位置和活性。测量同步S期期间DNA拷贝数的增加允许精确确定基因组复制。为了绘制起点位置，复制叉在其起始位点附近停止;因此，拷贝数富集仅限于起点。使用荧光激活细胞分选从异步培养物产生复制时序曲线。应用这种技术，我们发现，单倍体和二倍体细胞的复制模式是不可区分的，这表明这两种细胞类型使用相同的队列的起源具有相同的活动。最后，增加测序深度允许从指数增长的培养物直接测量复制动态。这是第一次这种方法，称为标记频率分析，已成功地应用于真核生物。这些数据为研究基因组生物学提供了高分辨率的资源和方法框架。
Eukaryotic genomes are replicated from multiple DNA replication origins. We present complementary deep sequencing approaches to measure origin location and activity in Saccharomyces cerevisiae. Measuring the increase in DNA copy number during a synchronous S-phase allowed the precise determination of genome replication. To map origin locations, replication forks were stalled close to their initiation sites; therefore, copy number enrichment was limited to origins. Replication timing profiles were generated from asynchronous cultures using fluorescence-activated cell sorting. Applying this technique we show that the replication profiles of haploid and diploid cells are indistinguishable, indicating that both cell types use the same cohort of origins with the same activities. Finally, increasing sequencing depth allowed the direct measure of replication dynamics from an exponentially growing culture. This is the first time this approach, called marker frequency analysis, has been successfully applied to a eukaryote. These data provide a high-resolution resource and methodological framework for studying genome biology.
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作者：
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影响因子：
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