【作者】 杨小林；

【导师】 喻大昭； 戚华雄；

【作者基本信息】 华中农业大学 ， 植物病理学， 2012， 博士

【摘要】 水稻稻瘟病是危害全球水稻生产最严重的病害之一,通过抗性品种防治稻瘟病的措施在当前被认为既经济有效又与环境相和谐,能有效地减少稻瘟病直接或间接所带来的损失。然而在实际生产中,含有单一抗性基因的抗性品种,长时期使用后会因为稻瘟病菌新的优势种群的出现而抗性渐失,因此,通过对稻瘟病菌群体致病型结构和遗传结构及其变异规律的分析；对抗性基因开展田间抗瘟性评价；同时利用不同的的水稻材料,对新的抗病基因进行探索等研究,可以为抗性品种的合理利用、克服抗病品种的感病化提供理论依据,从而长期、有效地控制该病害。基于中国鉴别品种和单基因品系以及rep-Pot2-PCR指纹技术,对湖北省稻瘟病重发区病菌群体的毒性结构及DNA指纹图谱进行了分析,结果表明,湖北省稻瘟病菌群体具有较丰富的毒性类型,根据对中国鉴别品种的致病性反应,2006-2010的单孢菌株可分成7群53个生理小种；根据对单基因品系的致病性反应,2009-2010年的菌株可分成72个致病类型。不同年份的稻瘟病菌群体在致病型组成上有差异,但在优势致病类型上没有明显的不同。在73%的指纹相似性水平上可将204菌株划分成112个不同的单元型和14个遗传谱系；14个谱系中没有出现绝对的优势谱系；不同年份稻瘟病菌群体遗传结构存在差异,但未发生明显的演化。DNA指纹分析的遗传谱系与生理小种(致病类型)之间呈复杂对应关系。湖北省稻瘟病重灾区稻瘟菌群体对抗性基因Pi-kh的毒性频率较低,表明Pi-kh基因在湖北省水稻稻作区有很好的利用价值。湖北省的恩施、宜昌、咸宁地区,一直是湖北省水稻稻瘟病的重发区和常发区。为了明确20个抗稻瘟病基因在湖北省的抗性水平及利用价值,选择恩施地区的金子坝和两河口、宜昌地区的远安县和咸宁地区的崇阳县作为水稻稻瘟病自然诱发接种试验点,对以丽江新团黑谷为轮回亲本培育而成的持有垂直抗性基因的20个水稻单基因系进行稻瘟病自然诱发接种。结果表明：抗性基因Pi-kh、Pi-9、Pi-20和Pi-z5对叶瘟有很好的抗性,而抗性基因Pi-a、Pi-t、Pi-ta、Pi-zt、Pi-12(t)对叶瘟的抗性不好。抗性基因Pi-kp、Pi-kh、Pi-5(t)、Pi-9和Pi-z5对穗颈瘟也有很好的抗性,而抗性基因Pi-t、Pi-ta, Pi-zt、Pi-12(t)、Pi-z对穗颈瘟的抗性不好。总结3年来20个垂直抗性基因在恩施、宜昌和咸宁的抗性情况,结果表明抗性基因Pi-kh、Pi-9和Pi-z5对稻叶瘟和抗稻穗颈瘟均有良好的抗性,可以作为湖北省水稻抗性基因或抗源基因合理使用,而抗性基因Pi-t、Pi-ta、Pi-z1、Pi-12(t)因对稻叶瘟和稻穗颈瘟的抗性较差,所以在推广使用时应谨慎。利用22个稻瘟病菌鉴别菌株对600份水稻品种(系)进行了抗瘟性测定,并对测定结果进行了聚类分析和品种(系)基因型推导。抗病性测定结果表明,600份品种(系)中有252份品种(系)对鉴别菌株表现出稳定的抗感反应,且这252份品种(系)对鉴别菌株的抗谱存在明显的多样性。根据供试品种(系)对鉴别菌株抗感反应型的聚类分析,可将252份品种(系)划分为15个组,各组的品种(系)对鉴别菌株的反应型和抗性频率存在明显差异。基因型推导结果表明,在252份品种(系)中,70份品种涉及14个抗瘟基因,181份品种(系)含有与已知的抗性基因都不相同的抗性因子。聚类分析结果与基因型推导结果不存在简单的一一对应关系。珍科、魔王谷等10份品种(系)可作为田间抗瘟性筛选的候选材料。以抗稻瘟病的云南省地方品种魔王谷(MWG)和感稻瘟病的湖北省地方品种鄂晚8号(EW8)为亲本材料,建立的双单倍体分离群体(DH)由96个株系组成,由此建立的分子遗传图谱包含120对SSR分子标记。通过菌株对亲本材料MWG/EW8不同的致病性反应,从22个菌株中筛选得到5个毒性不同的鉴别菌株,利用这5个鉴别菌株对DH系进行人工注射接种,考察该DH群体对稻瘟病的抗性。把DH系每一株系的病级数据输入到DH群体的分子标记数据库中,利用MAPMAKER/EXP3.0软件采用区间作图法对抗稻瘟病数量性状位点(QTL)进行检测,共鉴定出3个抗性QTL,均位于第6染色体长臂RM541附近,3个抗病QTL对抗病表型的贡献率介于7.7%-15.2%之间。3个抗病QTL均来源于抗病亲本MWG。更多还原

【Abstract】 Rice blast, caused by the fungal pathogen Magnaporthe oryzae, is the most economically important disease of rice in Hubei Province, where there are3×105hm2rice-growing fields. Although cultivars with resistance to rice blast have been released in Hubei, they had not been efficiently explored in the Hubei rice breeding programs. The major reason for the low efficiency is that the resistance is easily overcomed by the arisen pathogen lately. In order to provide scientific information to allow the formulation of viable strategies for blast resistance breeding programs in Hubei Province and to efficiently and durably control the disease, the study on current information on the pathotype spectrum of Magnaporhthe grisea population in the rice fields of Hubei Province, on evaluating performance of rice blast resistance genes, and identifing new gene by using various rice varieties, would be made in the dissertation.500single spore isolates in the rice-growing fields of Yuan’an in Hubei Province where rice blast seriously occurs in some years.500isolates were pathotyped against two sets of host differentials:Chinese host differentials and monogenic lines (MLs).72pathotypes with the nine host differentials which carried known resistance genes for blast resistance of the rice blast fungus Magnaporthe oryzae were observed, while7race groups53pathotypes were observed out of the same single spore isolates with Chinese host differentials which consists of three indica cultivars and four japonica cultivars. The spatial and temporal subpopulation pathotype structures of the fungus were also analyzed and the result revealed that there was no obvious change of the population among different years, and the prevalent pathotype hold the line. At the same time, DNA fingerprints hapotypes were determined. DNA fingerprints were separated into14lineages at73%genetic similarity level. And among the lineage, none of the most prevalent linelage was found. The population genetic structures of Magnaporthe oryzae were not distinctly different in different years. The assay also showed that there weren’t obvious and simple relationships between pathotypes and fingerprint groups. The gene Pi-kh had low susceptible frequency while resisting the rice blast fungus. The blast resistant gene Pi-kh can be well explored in blast resistance breeding programs in Hubei province.In order to evaluate performance of rice blast resistance genes in Hubei province,20monogenetic lines (MLs) derived from LTH were inoculated by nature in Jinziba and Lianghekou in Enshi district, Yuan’an in Yichang district and Chongyang district in Hubei Province, where blast occurs frequently and seriously in several years. The result revealed that genes Pi-kh, Pi-9, Pi-20and Pi-z5were incompatible to leaf blast while Pi-a, Pi-t, Pi-ta, Pi-z1,Pi-12(t) were compatible. Genes Pi-kp, Pi-kh,Pi-5(t), Pi-9and Pi were highly resistant to panicle blast while Pi-t, Pi-ta, Pi-zt, Pi-12(t, Pi-z were not resistant. Overall, there were genes Pi-kh, Pi-9and Pi-z5which had good resistance to rice blast including leaf blast and panicle blast can be used as rice blast resistance genes in Hubei rice breeding program, while it should be careful when the genes Pi-t, Pi-ta, Pi-z’, Pi-12(t were used as resistance genes in Hubei.Assay for resistance of600rice varieties(lines) to rice blast disease was accomplished using22differential isolates of Magnaporthe oryzae. Based on the result, clustering analysis and genotype postulation were completed by unweighted pair group method with arithmetic mean (UPGMA). Resistance test data showed that significant diversity of resistance spectrum to differential isolates was found among the total of252rice varieties(lines).These varieties(lines) were classified into15groups according to the reaction patterns of resistance or susceptibility to differential isolates. Tested varieties(lines) belonging to different groups displayed distinct variation for reaction patterns and resistance frequncy. Genotype postulation indicated that14Pi-genes were involved in70of252varieties(lines). Another181varieties harbored none of the tested Pi-genes, however, they all carried other Pi-genes. The result from clustering analysis was not corresponding to those from genotype postulation simply. Ten varieties(lines), such as Zhenke and Mowanggu, were able to be candidate materials for field resistance evalution.A doubled haploid (DH) segregation population, which was composed of96different varieties, derived from cross between Mowanggu, an famous resistance parent to rice blast from Yun’nan Province, and E’wan8, a susceptible parent from Hubei Province, was used to analyse quantitative trait locus (QTL) for resistance to rice blast fungal pathogen Magnaporthe oryzae. The graphical genotypes was investigated using120simple sequence repeat (SSR) markers distributed on the whole rice genome chromosomes.5isolates were screened among22isolates of Magnaporthe oryzae, which could create insusceptible/susceptible reaction on Muwanggu/E’wan8, respectively. Resistance to rice blast of the DH was monitored using the5isolates by injecting the conidial suspension into the DH lines. QTLs analysis using phenotype and molecular markers was performed in the DH segregation population through composite interval mapping with MAPMAKER/Exp3.0.3QTLs were found to have linkgage with SSR marker RM541on chromosome6with variance explained between7.7%-15.2%. The3resistance QTLs were all donated by Mowanggu.更多还原