Yu Guihong, Ma Hongxiang, Xu Zhang, Ren Lijian, Zhou Maoping, Lu Weizhong
Institute of Agro-Biological Genetics and Physiology, Jiangsu Academy of Agricultural Sciences, Nanjing, PR China.
J Appl Genet. 2004;45(1):17-25.
Wheat head blight caused mainly by Fusarium graminearum, is an important wheat disease, causing yield and quality losses. The breeding of resistant varieties is the key measure to control this disease, but the conventional breeding method is of low efficiency. The marker-assisted selection (MAS) can significantly improve the breeding efficiency. In this study, four RAPD (randomly amplified polymorphic DNA) markers linked to FHB resistance were obtained and one was cloned and sequenced. F7 recombinant inbred lines (RILs) were derived from the F1 of the cross Ning894037 (resistant)/Alondra (susceptible) by the single-seed descent method. Scab resistance of F7 RILs was evaluated in the greenhouse by injecting conidiospores into a central floret. Scab symptoms were evaluated on the 21st day after inoculation. Disease severity was assessed as the percentage of infected spikelets/spike. The F7 RIL population displayed a normal distribution, transgressive segregation and significant variation for FHB severity. DNA from resistant and susceptible parents was analyzed with 520 RAPD primers. Four markers (S1384-640,S1360-600, S1319-350,S1319-820) linked to FHB resistance were obtained. DNA of S1384-640 was recovered, subjected to re-amplification by using S1384 primer and the same protocol as for RAPD analysis and identified the rightness. The PCR product of S1384-640 was ligated into the pUCm-T vector, and cloned into fresh competent cells of Escherichia coli strain DH5alpha RAPD analysis showed that the inserts of the recombinant plasmids were DNA of S1384-640. The sequencing result showed that the cloned fragment was 648 bp.
小麦赤霉病主要由禾谷镰刀菌引起，是一种重要的小麦病害，会导致产量和品质损失。培育抗病品种是控制该病的关键措施，但传统育种方法效率低下。分子标记辅助选择（MAS）可显著提高育种效率。本研究获得了4个与赤霉病抗性相关的随机扩增多态性DNA（RAPD）标记，并对其中1个进行了克隆和测序。F7重组自交系（RILs）通过单粒传法从宁894037（抗病）/阿隆德拉（感病）杂交组合的F1代衍生而来。在温室中通过向中部小花注射分生孢子来评估F7 RILs的赤霉病抗性。接种后第21天评估赤霉病症状。病害严重程度以感染小穗数/穗的百分比来评估。F7 RIL群体在赤霉病严重程度上呈现正态分布、超亲分离和显著变异。用520条RAPD引物分析抗病和感病亲本的DNA。获得了4个与赤霉病抗性相关的标记（S1384 - 640、S1360 - 600、S1319 - 350、S1319 - 820）。回收S1384 - 640的DNA，使用S1384引物并按照与RAPD分析相同的方案进行重新扩增，以鉴定其正确性。将S1384 - 640的PCR产物连接到pUCm - T载体中，并克隆到大肠杆菌DH5α菌株的新鲜感受态细胞中。RAPD分析表明重组质粒的插入片段是S1384 - 640的DNA。测序结果显示克隆片段为648 bp。
