Plant Genome. 2017 Jul;10(2). doi: 10.3835/plantgenome2016.12.0123.
Eastern filbert blight (EFB), caused by the pyrenomycete (Peck) E. Müller, is a devastating disease of European hazelnut ( L) in the US Pacific Northwest. A dominant allele at a single locus from the obsolete pollenizer 'Gasaway' confers a high level of resistance to EFB. To identify the gene responsible for resistance, we initiated map-based cloning efforts in a population of 1488 seedlings that segregated for resistance. Chromosome walking was initiated using primers designed from eight previously identified random amplified polymorphic DNA markers linked to resistance. The bacterial artificial chromosome (BAC) library was screened using the primer pairs in a polymerase chain reaction-based pooling and subpooling strategy. Here, we report construction of a high-resolution genetic map and a physical map of the resistance region. Further, we sequenced BACs in the resistance region and identified and annotated the coding sequences. In seven contigs <1 cM from the resistance locus, 233 genes were predicted. The putative genes were compared with sequences in GenBank using a BLASTP search. Fifty-one markers were placed on the high-resolution genetic map, including markers newly developed from the BACs. Segregation in the mapping population placed the resistance locus in a single contig of three BACs (43F13, 66C22, and 85B7). Two of the putative genes are in the p-loop NTPase and F-box super-families localized in a 135-kb BAC, which have previously been shown to have disease-resistance properties. Further mapping, complementation, and expression tests of the genes in these BACs is essential to confirm which confer resistance to EFB.
东部榛枯萎病(EFB),由外囊菌(Peck)E. Müller引起,是美国太平洋西北地区欧洲榛的毁灭性病害。来自已废弃花粉品种“Gasaway”的单一基因座的显性等位基因赋予对 EFB 的高水平抗性。为了鉴定与抗性相关的基因,我们在一个分离出抗性的 1488 株幼苗群体中启动了基于图谱的克隆工作。利用先前与抗性相关的八个随机扩增多态性 DNA 标记设计的引物,启动了染色体步行。使用聚合酶链反应为基础的池化和亚池化策略,筛选了细菌人工染色体(BAC)文库。在这里,我们报告了抗性区域的高分辨率遗传图谱和物理图谱的构建。此外,我们对抗性区域的 BAC 进行了测序,并鉴定和注释了编码序列。在距离抗性位点<1cM 的七个 contig 中,预测了 233 个基因。使用 BLASTP 搜索,将推定基因与 GenBank 中的序列进行了比较。在高分辨率遗传图谱上放置了 51 个标记,包括从 BAC 新开发的标记。在作图群体中的分离将抗性位点定位在三个 BAC(43F13、66C22 和 85B7)的单个 contig 中。两个推定基因位于 p-loop NTPase 和 F-box 超家族中,定位于一个 135kb 的 BAC 中,以前已经证明它们具有抗病性。在这些 BAC 中对这些基因的进一步作图、互补和表达测试对于确认其对 EFB 的抗性至关重要。