【作者】 张迪迪；

【导师】 徐芳森；

【作者基本信息】 华中农业大学 ， 植物营养学， 2014， 博士

【摘要】 硼是植物生长发育的必需营养元素,在植物体内具有重要的生理功能。然而,土壤缺硼是世界范围内重要的农业生产问题,成为限制多种作物产量的主要因子。甘蓝型油菜是世界上主要的油料作物之一,是需硼较高的物种,且对缺硼敏感。但是,不同甘蓝型油菜品种对低硼胁迫存在极显著的基因型差异。本研究利用筛选出的硼高效(抗低硼胁迫)品种青油10号和硼低效(对低硼胁迫敏感)品种Westar10构建的QW DH群体,对甘蓝型油菜硼高效相关性状进行了QTL分析、同时构建了硼高效主效QTL的近等基因系,获得的主要结果如下：1.甘蓝型油菜硼高效相关性状QTL的检测利用420个SSR标记和153个AFLP标记构建了QW DH遗传连锁图谱,该图谱覆盖了1923.15cM的遗传距离,平均标记密度为0.30marker/cM,相邻分子间的平均距离为3.35cM。通过三次独立重复营养液培养试验调查QW DH群体及亲本材料在正常硼(25μM B)和低硼(0.25μM B)水平下的表型,选取根长增加量、地上部干重、根干重、地上部硼含量、根硼含量、地上部硼累积量、根硼累积量和硼效率系数共8个性状为硼高效相关性状。利用QW DH图谱对这8个硼高效相关性状进行QTL扫描,共检测到71个QTLs,其中,包括正常硼水平下30个QTLs,低硼水平下35个QTLs和硼效率系数6个QTLs,单个QTL的表型贡献率范围为4.58%-48.34%。所检测到的QTLs分布在QW DH遗传连锁图谱的18条连锁群上,其中在A3,A4,A5和C4连锁群上存在着QTL簇。位于A3连锁群58.8-82.5cM区间的QTL簇,QTL-cluster-A3a,包含低硼水平下调控根长增加量、地上部干重、根干重、地上部硼累积量、根中硼累积量和硼效率系数的QTLs,贡献率为14.80%-48.34%,被认为是控制甘蓝型油菜硼高效的主效QTL位点。2.甘蓝型油菜高密度遗传连锁图谱的构建及硼高效相关性状QTL的检测基于甘蓝型油菜60K SNP:芯片技术,在上述QW DH群体中获得11,754个SNP标记,分为1,346个SNP-bin。与上述QW DH遗传连锁图谱整合,获得一张包含1,398个标记的高密度遗传连锁图谱,覆盖遗传距离为2139.34cM,平均分子标记密度为0.65marker/cM,相邻分子间的平均距离为1.53cM。根据两图谱上共有的标记,高密度图谱与原图谱间具有较好的线性关系。利用高密度遗传连锁图谱重新检查硼高效相关性状的QTL,共获得75个QTLs,正常硼和低硼水平下分别为30和41个QTLs,硼效率系数4个QTLs。单个QTL的表型贡献率范围为7.26%-31.56%。其中,有34个QTLs可在两个图谱中重复检测到。另外,在A3,A4和C4连锁群上同样存在QTL簇的分布,这些QTL簇与先前QW DH图谱检测的QTL簇位于染色体的相同位置。3.甘蓝型油菜三个硼高效QTL簇近等基因系的构建根据QW DH群体的硼高效QTL检查结果,将QTL-cluster-A3a, QTL-cluster-A4和QTL-cluster-A5作为目标QTLs。以硼低效亲本Westar10为轮回亲本,通过连续不断回交,构建其近等基因系。通过每一回交世代单株抗低硼胁迫表型鉴定、高世代回交单株目标QTLs簇的前背景和后背景基因型分析,最终获得6个含有不同硼高效目标QTL簇的BC3F1单株自交发展的BC4F1株系。对这6个株系进行抗低硼表型鉴定,均表现出明显的抗缺硼表型分离。其中,以同时含有3个目标QTLs簇的BC4F1-3株系的表型分离最为明显。在含有硼高效主效QTL-cluster-A3a的BC4F1-2株系中,筛选到一个在QTL区间基因型为杂合、其它区间为Westar10纯和的单株,进一步通过自交获得该单株的BC4F2群体。在该群体中,筛选到含有不同目标QTL片段的染色体代换系,根据后代表型测验,将QTL-cluster-A3a锁定在A3连锁群的CNU384-BnGMS436区间。4.甘蓝型油菜地上部矿质元素QTL的分析调查QW DH群体在正常硼和低硼水平下的地上部矿质元素的含量。与正常硼相比,低硼降低了地上部K和Mn的含量,增加了P,Fe和Zn的含量,对于Ca,Mg和Cu的含量影响不大。矿质元素在不同的硼水平下存在着显著的正相关或负相关。利用QW DH遗传连锁图,对地上部矿质元素含量进行QTL分析。分别检测到控制甘蓝型油菜地上部大中量元素和微量元素的QTLs为28和27个,表型贡献率为10.44%-30.97%。控制不同矿质元素含量的QTLs在QW DH遗传连锁图谱上存在共定位的现象。更多还原

【Abstract】 Boron (B) is an essentical element for the growth and development of plants, which has the important physiological functions in plants. However, B deficiency in soils is a major agricultural problem worldwide, becoming a main limiting factor for the production of many crops. Oilseed rape (Brassica napus L.) is one main oil crop in the world, which is a high B demand species and is extremely sensitive to B deficiency. Significant genotypic vatiation was existed among the different B. napus cultivars in response to B deficiency. In this study, quantitative trait locus (QTL) for B-efficiency traits in B. napus was analyzed using the QW DH population derived from a cross between B-efficienct parent Qingyou10and B-inefficient parent Westar10, and near-isgenic lines (NILs) for major effect QTLs for B efficiency were also developped. The main results are as follows:1. QTL analysis for B-efficiency traits in B. napusThe QW DH genetic map was constructed using420simple sequence repeat (SSR) and153amplified fragment length polymorphism (AFLP) markers. The total length of this map was1923.15cM with an average marker density of0.30marker/cM and an average interval of3.35cM between adjacent markers. Phenotypes of QW DH population and their partental lines were investigated under normal B (25μM B) and low B (0.25μM B) levels by three independent solution culture trials. A total of eight traits were selected as the B-efficiency traits, including increment of primary root length (IPRL), shoot dry weight (SDW), root dry weight (RDW), shoot B concentration (SBC), root B concentration (RBC), shoot B accumulation (SBA), root B accumulation (RBA) and B efficiency coefficient (BEC). The QTLs for these eight traits were analyzed using the QW DH genetic map. A total of71QTLs were obstained, including30under normal B level,35under low B level and6for BEC, which explained single the phenotypic variation ranged from4.58%to48.34%. The QTLs distributed on eighteen linkage groups in QW DH genetic map. Moreover, the QTL clusters were observed on A3, A4, A5and C4linkage groups. QTL-cluster-A3a located in the interval of58.8-82.5cM on A3, including the QTLs for IPRL, SDW, RDW, SBA, RBA at low B level and BEC whose phenotypic varation ranged from14.80%-48.34%, was deemed as the major QTL for B efficiency of B. napus.2. Construction of high density genetic map and QTL mapping for B-efficiency traitsBased on the Brassica60K SNP BeadChip Array, a total of11,754single nucleotide polymorphism (SNP) markers were obstained in the QW DH population, classified into the1,346bins. The high density genetic map was constructed using these SNP-bins and the primary markers in the QW DH genetic map. The final high density genetic map contained1,398markers, covered2139.34cM, with an average marker density of0.65marker/cM and an average distance between adjacent markers of1.53cM. Linear relationship was observed between two genetic maps based on the commom marker. The QTL for B-efficiency traits was reanalyzed using the high density genetic map. In total,75QTLs were detected for associated with B efficiency. Of them,30QTLs were detected under normal B level,41QTLs were detected under low B level and4QTLs for BEC, respectively. Each QTL accounted for7.26%-31.56%of phenotypic variation. Among these QTLs,34were common detected across QW DH and high density genetic maps. Moreover, the QTL clusters on A3, A4and C4were also observed in high density genetic map, which located on the same chromosome regions with those observed in QW DH genetic map.3. Construction near isogenic lines of three QTL clusters for B efficiency in B. napusBased on the QTL results in QW DH population, QTL-cluster-A3a, QTL-cluster-A4and QTL-cluster-A5were selected as the target QTLs for construction the NIL. Backcross populations were constructed using B-inefficienct parent Westar10as recurrent parent. Every progenies were screened by the phenotypc investigation response to B deficiency and foreground and bachgroud analysis for high progenies, finlly, six BC4F1lines derived from a cross between BC3F1contained the different target QTLs and Westar10were used for investigation the phenotypic varation response to B deficiency. BC4F1.3, including three target QTLs simultaneously, showed the most widely variations among six lines. Substitution mapping revealed that the major QTL, QTL-cluster-A3a, was mapped in the interval between CNU384-BnGMS436.4. QTL analysis for mineral concentraions in shoot of B. napusThe mineral concentrations in shoot of QW DH population were investigated under normal B and low B levels. Compared with normal B, the concentrations of K and Mn were increased by B deficiency, however, the concentrations of P, Fe and Zn were decreased. The concentrations of Ca, Mg and Cu were not affected by B deficiency. Significant positive or negative relationships were observed among most of mineral concentrations. QTL for mineral concentrations was analyzed using QW DH genetic map. A total of28and27QTLs for macro-element concentrations and micro-element concentrations were detected, resceptively, explaining10.44%-30.97%of phenotypic variation. The co-locations of QTLs for different mineral concentrations were observed in QW DH genetic map.更多还原