Fluorescent Arabidopsis tetrads:: a visual assay for quickly developing large crossover and crossover interference data sets
Fluorescent Arabidopsis tetrads:: a visual assay for quickly developing large crossover and crossover interference data sets
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DOI：
10.1038/nprot.2007.491
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发表时间：
2008-01-01
期刊：
影响因子：
14.8
通讯作者：
Copenhaver, Gregory P.
中科院分区：
文献类型：
--
作者：
Berchowitz, Luke E.;Copenhaver, Gregory P.
在大多数生物体中，一个交叉（CO）事件抑制了另一个邻近事件的机会。用于描述这种现象的术语是“CO干扰”。在这里，我们描述了一个协议，用于快速生成大的数据集，适合在开花植物，拟南芥CO干扰分析。我们采用了一种视觉检测，利用转基因标记构建编码花粉表达的荧光蛋白的三种颜色的四重突变体的背景。在这种遗传背景下，雄性减数分裂产物-花粉粒-保持物理附着，从而有利于四分体分析。我们已经开发了一个映射标记插入的库，当它们交叉在一起时，可以创建相邻的间隔，可以快速地同时筛选CO。该分析系统能够检测和区分单CO以及双链、三链和四链双CO。我们还描述了如何分析由这种方法产生的数据。使用该方案在野生型和突变体背景中产生和评分双间隔将需要22 - 27周。
In most organisms, one crossover (CO) event inhibits the chances of another nearby event. The term used to describe this phenomenon is 'CO interference'. Here, we describe a protocol for quickly generating large data sets that are amenable to CO interference analysis in the flowering plant, Arabidopsis thaliana. We employ a visual assay that utilizes transgenic marker constructs encoding pollen-expressed fluorescent proteins of three colors in the quartet mutant background. In this genetic background, male meiotic products - the pollen grains - remain physically attached thereby facilitating tetrad analysis. We have developed a library of mapped marker insertions that, when crossed together, create adjacent intervals that can be rapidly and simultaneously screened for COs. This assay system is capable of detecting and differentiating single COs as well as two-, three- and four-strand double COs. We also describe how to analyze the data that are produced by this method. To generate and score a double interval in a wild-type and mutant background using this protocol will take 22-27 weeks.