Comparison of pear ‘La France’ and its bud mutant ‘Giant Laf’ (2010 expt, receptacle at 1 week before full bloom stage)

Bud mutation (bud sport) is mutation occurred in one branch in a tree and the mutant branch shows different phenotype from normal branches. Because bud mutation occurs in a part of plant body, genomic backgrounds of mutant branch and normal branches are the same, except some mutations. Therefore bud mutants are ideal material to identify key genes for important traits of crops. We studied ‘Giant Laf’, a bud mutant setting large fruit, which generated spontaneously in European pear ‘La France’ tree. In ‘Giant Laf’, increase of cell size and DNA reduplication occurred specifically in fruit flesh. The DNA reduplication in ‘Giant Laf’ was observed at 1 week before full bloom stage. This suggested that DNA reduplication observed in ‘Giant Laf’ fruit is not endoreduplication, but endomitosis or nuclear fusion. To identify genes expressed differentially between in ‘Giant Laf’ and in ‘La France’, microarray analysis was performed with RNA from receptacle (fruit flesh) at 1 week before full bloom stage. To isolate the receptacle, laser microdissection was applied. Genes encoding proteins localized in nucleus and cytoskeleton were up-regulated in ‘Giant Laf’. Among these genes, we found several genes which shared homology with previously described DNA reduplication related genes. These are candidates of responsible gene of ‘Giant Laf’ mutation. The gene expression profiles between European pear (Pyrus communis) ‘La France’ and its bud mutant ‘Giant Laf’ were compared. RNA was extracted from receptacle which was isolated from flower bud at 1 week before full bloom stage by using laser microdissection. Three biological replications were analyzed.

芽变（bud sport）是指发生在树木某一分枝上的突变事件，该突变分枝的表型与正常分枝存在显著差异。由于芽变仅发生于植物的部分组织中，突变分枝与正常分枝的基因组背景除特定突变位点外完全一致，因此芽变材料是鉴定作物重要性状关键基因的理想研究体系。本研究以大果芽变品种‘Giant Laf’为对象，该品种自发产生于欧洲梨‘La France’植株中。在‘Giant Laf’的果肉组织中，特异性出现了细胞体积增大与DNA复制倍增现象。该DNA复制倍增现象可在盛花期前1周被观测到，这表明‘Giant Laf’果实中的DNA复制倍增并非核内复制（endoreduplication），而是核内有丝分裂（endomitosis）或细胞核融合。为鉴定‘Giant Laf’与‘La France’间的差异表达基因，研究团队以盛花期前1周的花托（果肉）RNA为样本开展了微阵列（microarray）分析。为精准分离花托组织，研究采用了激光显微切割（laser microdissection）技术。在‘Giant Laf’中，定位于细胞核与细胞骨架的编码蛋白基因呈现上调表达趋势。其中，多个与已报道的DNA复制倍增相关基因具有同源性的基因被筛选获得，这些基因即为‘Giant Laf’芽变性状的候选功能基因。本研究对欧洲梨（*Pyrus communis*）‘La France’与其芽变品种‘Giant Laf’的基因表达谱进行了比较分析。以激光显微切割分离的盛花期前1周的花芽花托为材料提取总RNA，共设置3次生物学重复并完成分析。