Meiotic crossover hotspots and coldspots within Arabidopsis disease resistance genes. Meiotic crossover hotspots and coldspots within Arabidopsis disease resistance genes

To study recombination at the fine-scale we used high-throughput sequencing to analyse >1,000 crossovers within the RAC1 R gene hotspot. This revealed focused intragenic crossovers, overlapping exons encoding the TIR, NBS and LRR domains (RAC1 pollen typing sequencing). To analyse chromatin structure we performed micrococcal nuclease digestion of wild type (Col-0) chromatin and gel purified the resulting ~150 bp mononucleosomal DNA band. This DNA was used to generate a library and paired-end sequencing performed (MNase-seq)

为在精细尺度下研究重组事件，我们采用高通量测序（high-throughput sequencing）技术，对RAC1 R基因热点区域内的1000余个交叉互换事件进行了分析。该分析依托RAC1花粉分型测序（RAC1 pollen typing sequencing）完成，揭示了集中分布的基因内交叉互换事件，其位点与编码TIR结构域（TIR）、NBS结构域（NBS）及LRR结构域（LRR）的外显子区域相互重叠。为解析染色质结构，我们对野生型（Col-0）染色质开展了微球菌核酸酶（micrococcal nuclease）消化，并通过凝胶纯化得到了约150 bp的单核小体DNA条带。所获DNA被用于构建测序文库，并进行了双端测序（MNase-seq）。