全基因组关联分析解析水稻剑叶及单株产量的遗传基础

剑叶是水稻主要的源器官,剑叶大小与产量呈极显著的正相关,揭示剑叶大小和单株产量的遗传基础对于选育株型适宜的水稻高产品种具有重要意义。本研究以来源广泛、多样性丰富的1016份全球水稻核心种质资源为材料,在湖北荆州连续两年考察了种质资源的剑叶大小和单株产量。结果显示,剑叶长、宽、面积和单株产量在两年变异丰富,受环境影响大;剑叶长、宽和面积与单株产量相关性程度较低,说明这些性状的遗传基础不同。利用4.8M高密度SNP基因型数据进行全基因组关联分析,共定位到31个影响水稻剑叶相关性状和单株产量的QTL,其中1个QTL(qFLW4)在两年均定位到,3个位点同时影响多个性状(qFLW4和qFLA4、qFLL6和qFLA6、qFLL10.1和qFLA10)。结合候选区间单倍型分析和生物信息学分析,鉴定到7个候选基因和10个有利等位基因。研究结果为克隆水稻剑叶性状新基因和通过分子标记辅助选择配置不同剑叶相关基因塑造理想株型提供基因资源。     

显性E1基因背景下大豆开花和成熟期的全基因组关联分析

【目的】深入解析大豆开花期和成熟期的遗传基础,发掘控制性状的重要基因组区间,为分子标记辅助选择育种和新基因克隆提供依据。【方法】以224份携带显性E1基因的大豆种质为试验材料,在4个种植环境下调查表型数据,采用R-mrMLM软件包中的6种多位点关联分析模型对大豆开花和成熟期进行全基因组关联分析。【结果】共检测到91个开花期和83个成熟期QTNs（quantitative trait nucleotides）,其中6个开花期和10个成熟期QTNs能在多环境中检测到。这些环境稳定QTNs分布于15个区间大小在90~490 Kb的单倍型（基因组）内,并有6个基因组区间为本研究新检测到。【结论】研究表明显性E1基因背景下大豆开花和成熟期的QTN构成复杂,位于5号染色体39.52~40.01Mb等15个基因组区间是该群体中控制大豆开花期或成熟期的重要位点。     

黄瓜核心种质幼苗耐热性评价及GWAS分析

为研究黄瓜苗期耐热性及筛选耐热核心种质材料,本试验选取86份核心种质,在夏季利用日光温室,采用开关风口方式控制高温环境(50±4℃),对三叶一心的幼苗进行耐热性处理及鉴定。以幼苗受害症状划分热害等级,以相应的热害指数为指标进行黄瓜苗期耐热性评价。结合核心种质重测序结果,进行幼苗耐热性全基因组关联分析。结果表明,供试黄瓜核心种质耐热性差异显著,两次调查变异系数分别为21.9%和22.5%。利用热害指数为指标进行聚类分析,把86份黄瓜核心种质划分为四大类群。全基因组关联分析共检测到7个与苗期耐热性相关位点gHII4.1、gHII5.1、gHII5.2、gHII6.1、gHII7.1、gHII4.2、gHII6.2。其中位于4号染色体的gHII4.1和gHII4.2,与幼苗耐热性关联最大,在此区段预测到67个候选基因。     

水稻生殖发育期耐热性的分子遗传机制研究进展北大核心CSCD

水稻在生殖发育期对高温胁迫非常敏感,高温热害已经成为水稻生产最重要的限制因子之一。阐明水稻在生殖发育期耐热性的分子遗传机制,挖掘耐热关键基因,利用基因工程以及分子标记辅助选择的方法改良水稻品种的耐热性具有重要意义。从水稻生殖发育期耐热性的数量性状基因定位(QTL)、转录以及蛋白水平的高温应答、耐热转基因研究领域的进展进行了概述。     

水稻生殖发育期耐热性的分子遗传机制研究进展

水稻在生殖发育期对高温胁迫非常敏感,高温热害已经成为水稻生产最重要的限制因子之一。阐明水稻在生殖发育期耐热性的分子遗传机制,挖掘耐热关键基因,利用基因工程以及分子标记辅助选择的方法改良水稻品种的耐热性具有重要意义。从水稻生殖发育期耐热性的数量性状基因定位(QTL)、转录以及蛋白水平的高温应答、耐热转基因研究领域的进展进行了概述。     

水稻子粒硒含量的遗传及QTL检测

富硒功能水稻具有富硒、优质、保健、防病、安全等特点,已成为当前研究的热点。子粒硒含量的QTL定位对研究富硒功能水稻的遗传育种具有重要的意义。以籼稻亲本奉新红米和明恢100杂交的145个株系的F2群体构建遗传连锁图谱,图谱拟合92个SSR标记位点,覆盖水稻基因组2187.5 cM,标记间平均遗传距离为23.7 cM,占水稻全基因组的49.2%。采用复合区间作图法,对水稻子粒硒含量进行QTL分析,在第5染色体上共检测到2个新的水稻子粒硒的QTL,对表型变异的贡献率分别为6.39%、8.01%。     

水稻孕穗期耐热种质资源的初步筛选

在人工气候室设定极端高温条件下,对来源于不同国家和地区的58份处于温度敏感期的水稻品种耐热性进行鉴定,以结实率下降的百分率为指标,通过方差分析,对水稻种质资源的耐热性进行评价。结果表明,种质间的耐热性存在显著差异,筛选出极端耐热品种2份（IRAT118和冷水白）。     

普通菜豆镰孢菌枯萎病抗性种质资源筛选及全基因组关联分析

普通菜豆镰孢菌枯萎病是严重制约菜豆(Phaseolus vulgaris)产量的主要病害之一。采用下胚轴双孔注射法对601份普通菜豆种质资源进行枯萎病抗性鉴定, 共筛选出4份高抗材料。在此基础上, 基于分布在全基因组上的3 765 456个单核苷酸多态性(SNP)标记, 进行全基因组关联分析, 以P<1×10^-5为阈值。结果检测到57个显著关联的SNP位点, 分布于1、2、6、8和11号染色体上; 共获得8个显著关联区域, 其中位于1号染色体上的区域1包含SNP最多(48个), 最显著SNP P值为2.18E-07。在8个显著关联区域中, 共检测到186个基因, 其中157个基因有注释信息, 编码过氧化物酶、抗病蛋白、转录因子和蛋白激酶等。结合KEGG富集分析和序列同源性比对, 鉴定出9个候选基因可能与抗性相关。     

水稻糯性相关基因的全基因组关联分析

利用1 090 071个SNP标记, 对419份广西地方稻种资源核心种质的糯性进行全基因组关联分析。结果表明, 运用混合线性模型, 在P<4.72×10^-8 (4.72E-8)水平下, 检测到45个与糯性显著关联的SNP位点, 均位于第6号染色体上。通过筛选显著关联位点上、下游各150 kb区域内的基因, 共找到305个候选基因, 其中包含2个与淀粉合成酶相关的Wx和SSIIa基因; 同时在第6号染色体5.69-5.89 Mb区域发现4个与糯性显著关联的SNP位点, 该区域可能是影响水稻(Oryza sativa)糯性的重要候选区域。     

镉胁迫下甜、糯玉米开花期性状的全基因组关联分析

以183份甜、糯玉米自交系构成的关联群体为材料,在2种Cd污染水平下对甜、糯玉米开花期(DA,days to anthesis)、吐丝期(DS,days to silking)和开花吐丝间隔(ASI,anthesis silking interval)进行全基因组关联分析,以揭示重金属Cd胁迫对甜、糯玉米开花期的影响及调控基因。结果表明:不同Cd污染水平对甜、糯玉米DA和DS均产生影响,其中对DS的延长更为明显,从而导致ASI的增大。同时,甜玉米ASI平均值高于糯玉米ASI,由此表明甜玉米ASI对重金属Cd胁迫更为敏感。结合群体基因型和开花期相关性状表型的全基因组关联分析,分别筛选到3个开花期相关的SNP位点和6个吐丝期相关的SNP位点。根据其所对应的物理位置与Maize GDB和NCBI基因数据库进行比对,预测获得8个已知功能基因,其中包括在前人研究中与玉米开花期有关的候选基因。以上研究解析甜、糯玉米在重金属Cd污染水平下开花期及吐丝期的遗传规律,为今后开展甜、糯玉米分子标记辅助育种和Cd安全品种的选育提供参考。     

地方稻资源D43的开花期耐热特性研究

水稻在抽穗开花期对高温胁迫非常敏感,通过挖掘耐热资源,培育耐热水稻品种是应对高温热害最有效的方式。前期研究发现,地方稻资源D43在花期连续高温条件下能保持较高的结实率。本研究在大田和人工气候室不同高温处理下,分析了D43的开花时间与耐热性之间的相互关系。结果表明,高温能够使水稻的开花时间提前,D43表现出稳定的早花时特性,高温胁迫下的开花时间集中在8∶30~10∶00;在开花时间段恒定高温胁迫下,D43的结实率较低;但在大田高温和人工气候室模拟高温胁迫下,D43的开花时间避开了日高温段,从而表现出较高的结实率;花器官形态性状包括花药开裂率、柱头上的花粉附着数、花粉萌发数与结实率之间呈显著正相关,因此可用于评价水稻的花期耐热性。     

Quantitative trait loci associated with drought tolerance at reproductive stage in rice

Drought is a major constraint to rice (Oryza sativa) yield and its stability in rainfed and poorly irrigated environments. Identifying genomic regions influencing the response of yield and its components to water deficits will aid in our understanding of the genetics of drought tolerance and development of more drought tolerant cultivars. Quantitative trait loci (QTL) for grain yield and its components and other agronomic traits were identified using a subset of 154 doubled haploid lines derived from a cross between two rice cultivars, CT9993-510 to 1-M and IR62266-42 to 6-2. Drought stress treatments were managed by use of a line source sprinkler irrigation system, which provided a linearly decreasing level of irrigation coinciding with the sensitive reproductive growth stages. The research was conducted at the Ubon Rice Research Center, Ubon, Thailand. A total of 77 QTL were identified for grain yield and its components under varying levels of water stress. Out of the total of 77 QTL, the number of QTL per trait were: 7-grain yield (GY); 8-biological yield (BY); 6-harvest index (HI); 5-d to flowering after initiation of irrigation gradient (DFAIG); 10-total spikelet number (TSN); 7-percent spikelet sterility (PSS); 23-panicle number (PN); and 11-plant height (PH). The phenotypic variation explained by individual QTL ranged from 7.5% to 55.7%. Under well-watered conditions, we observed a high genetic association for BY, HI, DFAIG, PSS, TSN, PH, and GY. However, only BY and HI were found to be significantly associated with GY under drought treatments. QTL flanked by markers RG104 to RM231, EMP2_2 to RM127, and G2132 to RZ598 on chromosomes 3, 4, and 8 were associated with GY, HI, DFAIG, BY, PSS, and PN under drought treatments. The aggregate effects of these QTL on chromosomes 3, 4, and 8 resulted in higher grain yield. These QTL will be useful for rainfed rice improvement, and will also contribute to our understanding of the genetic control of GY under drought conditions at the sensitive reproductive stage. Close linkage or pleiotropy may be responsible for the coincidence of QTL detected in this experiment. Digenic interactions between QTL main effects for GY, BY, HI, and PSS were observed under irrigation treatments. Most (but not all) DH lines have the same response in measure of productivity when the intensity of water deficit was increased, but no QTL by irrigation treatment interaction was detected. The identification of genomic regions associated with GY and its components under drought stress will be useful for marker-based approaches to improve GY and its stability for farmers in drought-prone rice environments.     

水稻红莲型CMS育性恢复QTL分析

红莲型CMS是在我国杂交水稻生产中被广泛利用的雄性不育细胞质之一。为了同时定位红莲型CMS育性恢复主效和微效QTL,利用红莲型CMS不育系粤泰A(YTA)与“Lemont／特青”RIL群体测交,结合1张含有198个DNA分子标记的高密度遗传图谱,对测交F1群体的小穗育性和花粉育性进行复合区间作图。在对YTA的育性恢复性方面,该。RIL群体的2个亲本之间具有明显差异,特青的恢复性较强,其测交F1的小穗育性和花粉育性分别为72％和51％;而Lemont测交F1的小穗育性和花粉育性分别为32％和9％。复合区间作图定位到4个育性恢复QTL,分别位于水稻第1、2和10号染色体上,单个QTL的贡献率在5％～24％之间。其中,除1个QTL的增效基因来源于Lemont外,其余3个QTL的增效基因均来源于特青。效应最大的QTL为qRF-10-1,该QTL位于10号染色体RM258-C16标记区间,对小穗育性表型变异的贡献率为24％,对花粉育性的贡献率为17％,且该QTL被检测到的LOD值显著较高,因此是1个主效QTL,其增效基因来源于特青。除了主效QTLqRF-10-1外,其它3个QTL对性状的贡献率均在10％以下(5％～8％)。由此表明,该RIL群体对红莲型CMS的育性恢复由1个主效QTL控制,并受其它几个微效QTL的影响。该QTL定位结果与小穗育性在测交F1群体中呈连续的双峰分布的结果相一致。与主效QTL qRF-10-1紧密连锁的SSR标记为RM258,该主效QTL可作为分子标记辅助育种的操作目标之一,用于杂交稻分子育种中培育红莲型CMS的强恢复系。     

Genetic analysis for drought resistance of rice at reproductive stage in field with different types of soil

Drought resistance of rice is a complex trait and is mainly determined by mechanisms of drought avoidance and drought tolerance. The present study was conducted to characterize the genetic basis of drought resistance at reproductive stage in field by analyzing the QTLs for drought response index (DRI, normalized by potential yield and flowering time), relative yield, relative spikelet fertility, and four traits of plant water status and their relationships with root traits using a recombinant inbred population derived from a cross between an indica rice and upland rice. A total of 39 QTLs for these traits were detected with individual QTL explained 5.1–32.1% of phenotypic variation. Only two QTLs for plant water status were commonly detected in two environments, suggesting different mechanisms might exist in two types of soil conditions. DRI has no correlation with potential yield and flowering time under control, suggesting that it can be used as a good drought resistance index in field conditions. The co-location of QTLs for canopy temperature and delaying in flowering time suggested a usefulness of these two traits as indexes in drought resistance screening. Correlation and QTL congruence between root traits and putative drought tolerance traits revealed that drought avoidance (via thick and deep root traits) was the main genetic basis of drought resistance in sandy soil condition, while drought tolerance may play more role in the genetic basis of drought resistance in paddy soil condition. Therefore, both drought mechanisms and soil textures must be considered in the improvement of drought resistance at reproductive stage in rice.     

Pollen germination and in vivo fertilization in response to high‐temperature during flowering in hybrid and inbred rice

High‐temperature during flowering in rice causes spikelet sterility and is a major threat to rice productivity in tropical and subtropical regions, where hybrid rice development is increasingly contributing to sustain food security. However, the sensitivity of hybrids to increasing temperature and physiological responses in terms of dynamic fertilization processes is unknown. To address these questions, several promising hybrids and inbreds were exposed to control temperature and high day‐time temperature (HDT) in Experiment 1, and hybrids having contrasting heat tolerance were selected for Experiment 2 for further physiological investigation under HDT and high‐night‐time‐temperature treatments. The day‐time temperature played a dominant role in determining spikelet fertility compared with the night‐time temperature. HDT significantly induced spikelet sterility in tested hybrids, and hybrids had higher heat susceptibility than the high‐yielding inbred varieties. Poor pollen germination was strongly associated with sterility under high‐temperature. Our novel observations capturing the series of dynamic fertilization processes demonstrated that pollen tubes not reaching the viable embryo sac was the major cause for spikelet sterility under heat exposure. Our findings highlight the urgent need to improve heat tolerance in hybrids and incorporating early‐morning flowering as a promising trait for mitigating HDT stress impact at flowering.     

High temperature stress and spikelet fertility in rice (Oryza sativa L.)

In future climates, greater heat tolerance at anthesis will be required in rice. The effect of high temperature at anthesis on spikelet fertility was studied on IR64 (lowland indica) and Azucena (upland japonica) at 29.6 degrees C (control), 33.7 degrees C, and 36.2 degrees C tissue temperatures. The objectives of the study were to: (i) determine the effect of temperature on flowering pattern; (ii) examine the effect of time of day of spikelet anthesis relative to a high temperature episode on spikelet fertility; and (iii) study the interactions between duration of exposure and temperature on spikelet fertility. Plants were grown at 30/24 degrees C day/night temperature in a greenhouse and transferred to growth cabinets for the temperature treatments. Individual spikelets were marked with paint to relate fertility to the time of exposure to different temperatures and durations. In both genotypes the pattern of flowering was similar, and peak anthesis occurred between 10.30 h and 11.30 h at 29.2 degrees C, and about 45 min earlier at 36.2 degrees C. In IR64, high temperature increased the number of spikelets reaching anthesis, whereas in Azucena numbers were reduced. In both genotypesor=33.7 degrees C at anthesis caused sterility. In IR64, there was no interaction between temperature and duration of exposure, and spikelet fertility was reduced by about 7% per degrees C>29.6 degrees C. In Azucena there was a significant interaction and spikelet fertility was reduced by 2.4% degrees Cd-1 above a threshold of 33 degrees C. Marking individual spikelets is an effective method to phenotype genotypes and lines for heat tolerance that removes any apparent tolerance due to temporal escape.     

Physiological and biochemical characterization of NERICA‐L‐44: a novel source of heat tolerance at the vegetative and reproductive stages in rice

The predicted increase in the frequency and magnitude of extreme heat spikes under future climate can reduce rice yields significantly. Rice sensitivity to high temperatures during the reproductive stage is well documented while the same during the vegetative stage is more speculative. Hence, to identify and characterize novel heat‐tolerant donors for both the vegetative and reproductive stages, 71 rice accessions, including approximately 75% New Rice for Africa (NERICAs), were phenotyped across field experiments during summer seasons in Delhi, India, and in a controlled environment study at International Rice Research Institute , Philippines. NERICA‐L‐44 (NL‐44) recorded high seedling survival (52%) and superior growth and greater reproductive success exposed to 42.2°C (sd ± 2.3) under field conditions. NL‐44 and the heat‐tolerant check N22 consistently displayed lower membrane damage and higher antioxidant enzymes activity across leaves and spikelets. NL‐44 recorded 50–60% spikelet fertility, while N22 recorded 67–79% under controlled environment temperature of 38°C (sd ±1.17), although both had about 87% fertility under extremely hot field conditions. N22 and NL‐44, exposed to heat stress (38°C), had similar pollen germination percent and number of pollen tubes reaching the ovary. NL‐44 maintained low hydrogen peroxide production and non‐photochemical quenching (NPQ) with high photosynthesis while N22 avoided photosystem II damage through high NPQ under high‐temperature stress. NL‐44 with its reproductive stage resilience to extreme heat stress, better antioxidant scavenging ability in both vegetative tissue and spikelets and superior yield and grain quality is identified as a novel donor for increasing heat tolerance at both the vegetative and reproductive stages in rice.     

Field level evaluation of rice introgression lines for heat tolerance and validation of markers linked to spikelet fertility

Rice lines derived from wild species and mutants can serve as a good resource for favorable alleles for heat tolerance. In all, 48 stable lines including 17 KMR3/O. rufipogon introgression lines (KMR3 ILs), 15 Swarna/O. nivara ILs (Swarna ILs) along with their parents, Nagina 22 (N22) and its 4 EMS induced mutants and 7 varieties were evaluated for heat tolerance under irrigated conditions under field in two seasons, wet season 2012 using poly cover house method and dry season 2013 using late sown method. Spikelet fertility (SF), yield per plant (YP) and heat susceptibility index (HSI) for these two traits were considered as criteria to assess heat tolerance compared to control. Four KMR3 ILs and eight Swarna ILs were identified as heat tolerant based on SF and YP and their HSIs in both wet and dry seasons. S-65 and S-70 showed low SF and high YP consistently in response to heat in both seasons. We provide evidence that SF alone may not be the best criterion to assess heat tolerance and including YP is important as lines with low SF but high YP and vice versa were identified under heat stress. Out of 49 SSR markers linked to spikelet fertility, 18 were validated for five traits. RM229 in wet season and RM430 and RM210 in dry season were significantly associated with both SF and its HSI under heat stress. RM430 was also significantly associated with both YP and its HSI in dry season. Thirty two candidate genes were identified close to nine markers associated with traits under heat stress.     

两个水稻骨干恢复系重要农艺性状的遗传基础研究

水稻骨干恢复系是指在杂交稻育种中广泛应用的一类恢复系。探明骨干恢复系的遗传基础,发掘其重要农艺性状基因/QTL,对分子标记辅助选择水稻恢复系育种具有重要应用价值。本研究以生产上广泛应用的三系骨干恢复系成恢727和两系骨干恢复系9311为亲本,培育了具有250个系的重组自交系群体。分别在2015年三亚和2016年合肥两个环境下进行了9个重要农艺性状表型和SSR分子标记基因型鉴定,用SAS9.2分析表型数据,用QTL Ici Mapping v4.1进行QTL定位分析。在三亚和合肥两个环境下共检测到39个QTL,三亚检测到16个,分布于第1、2、4、7、8、10、11和12染色体上;合肥检测24个,分布于第1、2、3、7、8、9、10和12染色体上。其中qPH1-1在三亚和合肥两个环境下都能检测到,加性效应分别为-1.75和-2.46。在检测到的39个QTL中,有24个QTL的增效等位基因来自恢复系成恢727,15个QTL的增效等位基因来自9311。共计有26个QTL曾被前人定位,13个属于尚未见文献报道的新QTL。另外,在RM279~RM521、RM336~RM3534、RM25~RM547、RM553~RM160、RM222~RM271区段内检测到5个多效性QTL位点。其中RM25~RM547位点与已经克隆的基因Ghd8位置相近。RM553~RM160位点是一个新的多效性位点,分别控制每穗实粒数、单株产量和结实率,而且效应和表型变异贡献率都较大。其余3个位点在前人的研究中分别有所报道,但其多效性则是在本研究中首次发现。在本研究新发掘到的QTL中,控制穗数的QTL qPN12-1,控制穗长的QTL qPL1-2和qPL10-1,控制总粒数的QTL qSNP2-1和qSNP10-1,控制结实率的QTL qSF3-1,控制千粒重QTL qTGW7-1和控制产量的QTL qGY1-1效应均比较大,解释的表型遗传变异比例也较高。本研究的结果将会为相关性状QTL的精细定位、克隆和育种应用奠定基础。     

基于染色体置换系的普通野生稻耐盐性QTL定位

本研究分别利用SSR标记、InDel标记以及简化基因组测序技术对一套以普通野生稻为供体亲本,9311为受体亲本的染色体置换系进行基因型鉴定,并通过对其不同生育期的耐盐性鉴定,共发掘2个与发芽期耐盐相关的QTL,13个与苗期耐盐性相关的QTL。其中与苗期存活率相关的QTL qSSR5.1、苗期耐盐等级相关的QTL qSSG5.1均被定位于同一位点,该位点对苗期存活率、苗期耐盐等级均具有增效作用,贡献率分别为6.36%、8.13%。在此QTL内包含与非生物胁迫相关基因OsDi19-1。经序列比对发现,OsDi19-1启动子区域在两亲本间存在较大差异,且受到盐胁迫时该基因表达量上升。同时,鉴定出水稻芽期耐盐的优异种质资源CSSL72、苗期耐盐的优异种质资源CSSL23、CSSL153,为水稻育种中耐盐性的改良提供了新的种质资源。