Dracatos Peter M, Cogan Noel O I, Sawbridge Timothy I, Gendall Anthony R, Smith Kevin F, Spangenberg German C, Forster John W
Department of Primary Industries, Biosciences Research Division, Victorian AgriBiosciences Centre, La Trobe University Research and Development Park, Bundoora, Victoria, Australia.
BMC Plant Biol. 2009 May 19;9:62. doi: 10.1186/1471-2229-9-62.
Qualitative pathogen resistance in both dicotyledenous and monocotyledonous plants has been attributed to the action of resistance (R) genes, including those encoding nucleotide binding site--leucine rich repeat (NBS-LRR) proteins and receptor-like kinase enzymes. This study describes the large-scale isolation and characterisation of candidate R genes from perennial ryegrass. The analysis was based on the availability of an expressed sequence tag (EST) resource and a functionally-integrated bioinformatics database.
Amplification of R gene sequences was performed using template EST data and information from orthologous candidate using a degenerate consensus PCR approach. A total of 102 unique partial R genes were cloned, sequenced and functionally annotated. Analysis of motif structure and R gene phylogeny demonstrated that Lolium R genes cluster with putative ortholoci, and evolved from common ancestral origins. Single nucleotide polymorphisms (SNPs) predicted through resequencing of amplicons from the parental genotypes of a genetic mapping family were validated, and 26 distinct R gene loci were assigned to multiple genetic maps. Clusters of largely non-related NBS-LRR genes were located at multiple distinct genomic locations and were commonly found in close proximity to previously mapped defence response (DR) genes. A comparative genomics analysis revealed the co-location of several candidate R genes with disease resistance quantitative trait loci (QTLs).
This study is the most comprehensive analysis to date of qualitative disease resistance candidate genes in perennial ryegrass. SNPs identified within candidate genes provide a valuable resource for mapping in various ryegrass pair cross-derived populations and further germplasm analysis using association genetics. In parallel with the use of specific pathogen virulence races, such resources provide the means to identify gene-for-gene mechanisms for multiple host pathogen-interactions and ultimately to obtain durable field-based resistance.
双子叶植物和单子叶植物中的定性病原体抗性归因于抗性（R）基因的作用，包括那些编码核苷酸结合位点-富含亮氨酸重复序列（NBS-LRR）蛋白和类受体激酶的基因。本研究描述了从多年生黑麦草中大规模分离和鉴定候选R基因。该分析基于表达序列标签（EST）资源和功能整合的生物信息学数据库。
使用简并共有PCR方法，利用模板EST数据和直系同源候选基因的信息进行R基因序列扩增。共克隆、测序并进行功能注释了102个独特的部分R基因。基序结构和R基因系统发育分析表明，黑麦草R基因与推定的直系同源位点聚类，并起源于共同的祖先。通过对遗传作图家系亲本基因型扩增子的重测序预测的单核苷酸多态性（SNP）得到验证，26个不同的R基因位点被定位到多个遗传图谱上。大量不相关的NBS-LRR基因簇位于多个不同的基因组位置，并且通常与先前定位的防御反应（DR）基因紧密相邻。比较基因组学分析揭示了几个候选R基因与抗病数量性状位点（QTL）的共定位。
本研究是迄今为止对多年生黑麦草定性抗病候选基因最全面的分析。在候选基因中鉴定出的SNP为在各种黑麦草双杂交衍生群体中进行作图以及使用关联遗传学进行进一步的种质分析提供了宝贵资源。与使用特定病原体毒力小种并行，这些资源提供了识别多种宿主病原体相互作用的基因对基因机制并最终获得持久田间抗性的方法。
