【作者】 李广贤；

【导师】 李斯深； 姚方印；

【作者基本信息】 山东农业大学 ， 作物遗传育种， 2014， 博士

【摘要】 抽穗期决定水稻品种适宜的种植地区和种植季节，影响水稻的稳产和高产，是重要的水稻育种目标之一。探讨水稻抽穗期的遗传本质，发掘和鉴定水稻抽穗期基因并分析其光温反应特性，对于对指导育种实践和品种推广均具有重要意义。本研究以遗传背景一致的染色体单片段代换系为材料，鉴定和定位水稻抽穗期QTL，并分析它们之间的互作关系，以期获得准确可靠的研究结果，为抽穗期QTL克隆和分子标记辅助育种提供的理论依据。本研究获得以下主要结果：（1）次级单片段代换系的创建。通过SSR标记选择，从初级单片段代换系之间的杂交分离群体S1×S2， S1×S3，S1×S4和S1×S5中获得了20个纯合的次级单片段代换系，其中8个代换系的代换片段位于第3染色体且来源于同一供体，另外12个代换系的代换片段位于第6染色体，虽然代换片段供体来源不同，但具有相同或相近的代换区间。筛选获得的次级单片段代换系的代换片段长度相对于初级单片段代换系明显缩短。（2）利用单片段代换系鉴定水稻抽穗期QTL。自然长日照条件下，有20个单片段代换系的代换片段被鉴定出带有抽穗期QTL。经QTL代换作图分析，共定位出两个抽穗期QTL，其中qHD3位于第3染色体短臂末端，qHD6位于第6染色体短臂中部。研究还发现qHD3和qHD6都具有多种表型效应，其中qHD3早抽穗等位基因不仅促进抽穗，还降低株高、穗粒数和千粒重，而qHD6早抽穗等位基因则仅缩短抽穗期和降低株高，对产量性状没有显著的影响。（3）qHD3精细定位及候选基因分析。应用杂合的代换系分离群体将qHD3定位于第3染色体短臂SSR标记RM14314和RM569之间。通过扩大分离群体，最终将qHD3精细定位RM14314和RM14320之间到约62.4Kb的染色体区间。在qHD3所在的染色体区间存在一个MADS-box转录因子OsMADS50，该基因具有调控水稻抽穗开花的功能。基因序列分析发现OsMADS50的编码区存在一处单碱基转换，内含子序列存在两处变异，其中第二内含子上存在一处20个碱基的缺失/插入，第六内含子存在一处单碱基转换。在携带不同qHD3等位基因的单片段代换系和受体之间未发现OsMADS50转录水平的表达差异，说明上述单片段代换系和受体的抽穗期差异不是通过改变OsMADS50转录水平的表达量来调控的。（4）qHD6精细定位及候选基因分析。应用携带不同qHD6等位基因的单片段代换系之间的杂交分离群体将另一个抽穗期基因qHD6定位于第6染色体的短臂中部，qHD6介于SSR标记RM587和RM204之间，两标记之间的物理距离为876.6Kb，在此染色体区域已有两个水稻抽穗期基因（RFT1和Hd3a）被克隆。基因序列和转录水平的表达分析都发现RFT1和Hd3a在两个单片段代换系亲本之间存在差异，所以目前不能确定哪个基因是qHD6的目的基因。（5）qHD3和qHD6互作。分离群体和双基因聚合系分析都表明qHD3和qHD6之间存在显著的互作关系，其中早抽穗的qHD3等位基因对qHD6呈上位性。更多还原

【Abstract】 Heading date of rice (Oryza sativa L.) is a key agronomic trait for adaption to cultivatedareas and seasons in order to attain an ideal grain yield, and it is also one of the leadingobjectives of rice improvement programs. Studies on genetic regulatory mechanism forheading date are useful for rice breeding and productive practice. The objectives of thisstudy were to detect QTLs for heading date and analyze their interaction effects using singlesegment substitution lines (SSSLs) with consistent genetic background. This result for QTLanalysis will be more accurate and useful for QTL cloning and molecular marker assistedbreeding. The main results were following:(1) Development of secondary SSSLs (S-SSSLs)Twenty homozygous S-SSSLs were obtained from segregation populations derived fromfour crosses between primary SSSLs (P-SSSLs): S1×S2， S1×S3，S1×S4and S1×S5.Among them, eight S-SSSLs contained the substituted segments on the short arm ofchromosome3derived from the same donor ‘Lemont’ and others contained substitutedsegments on the short arm of chromosome6derived from four different donors,‘Tetep’,‘BG367’,‘IR58025B’ and ‘Lemont’, respectively. The length of substituted segments inS-SSSL had been greatly shortened compared with the corresponding P-SSSL.(2) QTL identification with SSSLsTwo QTLs, qHD3and qHD6on the short arm of chromosome3and the short arm ofchromosome6, respectively, were identified with twenty SSSLs under natural long-day(NLD). Both qHD3and qHD6had pleiotropic effects. qHD3early allele not only promotedheading, but also decreased plant height and the number of grains main panicle and affectedgrain shape and weight. qHD6affected heading date and plant height, but had no significanteffect on the other agronomic traits.(3) Fine mapping of qHD3and candidate gene analysisWith substitution lines segregating population, qHD3was located between SSR markersRM14314and RM14320, the distance between the two markers was62.4Kb. Based oncandidate gene analysis, we found a transcription factor, MADS-box gene OsMADS50(heading date gene) was in qHD3target chromosomal region. Sequence alignment ofOsMADS50derived from SSSL with qHD3early heading allele and recipient HJX74 showed that the code region sequence of OsMADS50existed a nucleotide substitution andits intron sequence existed two variants,20nucleotides delete/insert and a nucleotidesubstitution. However, we did not found the differential expression at transcription level forOsMADS50from SSSL with qHD3early heading allele and recipient HJX74. This resultsuggested that the phenotypic variation of SSSLs with different qHD3alleles is not relatedto the expression of OsMADS50at transcription level.(4) Fine mapping of qHD6and candidate gene analysisWith a F2segregating population derived from the cross between SSSLs which containeddifferent qHD6allele, qHD6was located on the short arm of chromosome6and betweenSSR markers RM587and RM204, the distance between the two markers was876.6Kb. Inthis chromosomal region existed two heading date gene, RFT1and Hd3a. Both RFT1andHd3a showed variations in gene sequence and expression at transcription level betweenSSSLs with different qHD6allele. Therefore, it is still unknown which gene was the targetgene of qHD6.(5) Analysis for interaction between qHD3and qHD6Analysis of double segments pyramid lines (DSPLs) and F2populations showed theexistence of epistatic interactions between qHD3and qHD6. Furthermore, qHD3wasepistatic to qHD6under NLD conditions.更多还原