Boyko Alexander, Filkowski Jody, Hudson Darryl, Kovalchuk Igor
Department of Biological Sciences, University of Lethbridge, Lethbridge, Alta., Canada T1K 3M4.
Mutat Res. 2006 Mar 20;595(1-2):145-55. doi: 10.1016/j.mrfmmm.2005.11.009.
In this paper we analysed the genome stability of various Arabidopsis thaliana plant organs using a transgenic recombination system. The system was based on two copies of non-functional GUS (lines #651 and #11) or LUC (line #15D8) reporter genes serving as a recombination substrate. Both reporter assays showed that recombination in flowers or stems were rare events. Most of the recombination sectors were found in leaves and roots, with leaves having over 2-fold greater number of the recombination events per single cell genome as compared to roots. The recombination events per single genome were 9.7-fold more frequent on the lateral half of the leaves than on the medial halves. This correlated with a 2.5-fold higher metabolic activity in the energy source (lateral) versus energy sink (medial) of leaves. Higher metabolic activity was paralleled by a higher anthocyanin production in lateral halves. The level of double strand break (DSB) occurrence was also different among plant organs; the highest level was observed in roots and the lowest in leaves. High level of DSBs strongly positively correlated with the activity of the key repair enzymes, AtKU70 and AtRAD51. The ratio of AtRAD51 to AtKU70 expression was the highest in leaves, supporting the more active involvement of homologous recombination pathway in the repair of DSBs in this organ. Western blot analysis confirmed the real time PCR expression data for AtKU70 gene.
在本文中,我们使用转基因重组系统分析了拟南芥不同植物器官的基因组稳定性。该系统基于两个无功能的GUS(#651和#11系)或LUC(#15D8系)报告基因拷贝作为重组底物。两种报告基因检测均表明,花或茎中的重组是罕见事件。大多数重组区域出现在叶和根中,与根相比,叶中每个单细胞基因组的重组事件数量多出两倍以上。每个单基因组的重组事件在叶的外侧半部比内侧半部频繁9.7倍。这与叶的能量源(外侧)相对于能量库(内侧)中高2.5倍的代谢活性相关。较高的代谢活性伴随着外侧半部较高的花青素产量。双链断裂(DSB)的发生水平在不同植物器官中也有所不同;根中观察到的水平最高,叶中最低。高水平的DSB与关键修复酶AtKU70和AtRAD51的活性呈强烈正相关。AtRAD51与AtKU70表达的比率在叶中最高,这支持同源重组途径在该器官DSB修复中更积极的参与。蛋白质印迹分析证实了AtKU70基因的实时PCR表达数据。