【作者】 杨进文；

【导师】 孙黛珍；

【作者基本信息】 山西农业大学 ， 作物遗传育种， 2017， 博士

【摘要】 我国是氮肥消费大国,同时氮肥利用率一直低于世界平均水平。在总体施氮量不增加的前提下,维持小麦高产必须提高小麦品种的氮素利用效率。氮高效基因型筛选是提高氮肥利用效率、减少环境污染的主要途径。本研究以16个小麦品种为试材,利用主成分分析和相关性分析筛选能确切评价氮素利用率的指标,然后以这些指标结合产量性状进行聚类分析筛选出氮素利用率不同的品种,并且对筛选的品种进行生理生化特性分析,揭示氮素积累转运生理基础;利用DH群体(旱选10号×鲁麦14)对灌浆期间小麦各主要器官氮素含量及氮素积累转运相关性状进行QTL分析,寻找稳定表达的QTL分布热点,为今后氮素高效利用小麦品种的选育提供参考。主要研究结果如下:1.以16个小麦品种为试材,测定了14个氮素积累转运相关性状,然后,选取变异系数较大的指标性状,再通过主成分分析和遗传相关分析确定了8个与氮素积累转运密切相关的指标性状,即花后氮素积累量、花前氮素转运量、花前氮素转运贡献率、花后氮素转运量、花后氮素转运率、花后氮素转运贡献率、籽粒氮素生产效率,氮素干物质生产效率。以这8个指标性状以及产量相关性状进行综合聚类,将16个品种分为3类,第一类为氮高效型品种包括晋麦54、晋麦66、FRFSCD共3个品种,第三类为氮低效型,包括晋麦72、泰农18、晋麦73及晋麦61共4个品种,第二类为中间类型,包括其余9个品种。2.从每一类型中选取2个品种共6个品种,进行氮素积累转运相关酶活性测定,两种水分条件下可溶性蛋白质、谷氨酰胺合成酶、谷氨酸合成酶、谷氨酰胺转移酶和天冬氨酸激酶在花后呈下降趋势,蛋白水解酶活性呈上升趋势;两种水分条件下品种间各期酶活的差距大小有所不同。灌浆期间,氮高型品种晋麦54、泰农66的酶活性一直高于氮低效和中间类型品种晋麦73、泰麦269、晋麦61、泰农18。对不同品种各种酶活性与氮素积累转运相关性状进行相关性分析,发现灌浆期不同时期大多数代谢关键酶活性与氮素籽粒生产效率、氮素干物质生产效率、花后氮素转运量、花后氮素转运效率、花后对籽粒氮积累的贡献率皆呈显著或极显著的遗传正相关,与花前对籽粒氮积累的贡献率、花前氮素转运量呈显著或极显著的遗传负相关。并且遗传相关系数高于表型相关系数,表明这6种氮素积累转运相关酶基因的表达确实影响着小麦氮素的积累与转运。3.两种水分条件下,利用非条件与条件QTL分析方法检测花后不同发育阶段叶片、茎秆、穗轴、籽粒和植株氮素含量QTL,发现控制叶片氮素含量的加性QTL分别为10个与7个,对表型变异的贡献率为5.18%～23.8%,上位性QTL分别是18对与11对,对表型变异的贡献率为0.50%～43.76%;控制茎秆氮素含量的加性QTL分别为12个与4个,对表型变异的贡献率为6.72%～19.67%,上位性QTL分别是19对与7对,对表型变异的贡献率为0.88%～33.63%;控制穗轴氮素含量的加性QTL分别是5个与4个,对表型变异的贡献率为8.65%～17.56%;上位性QTL分别是9对与9对,对表型变异的贡献率为0.81%～38.72%;控制籽粒氮素含量的加性QTL分别是14个与5个,对表型变异的贡献率为6.08%～27.44%;上位性QTL分别是9对与5对,对表型变异的贡献率为0.40%～30.34%;控制植株氮素含量的加性QTL分别是15个与11个,对表型变异的贡献率为6.07%～23.06%;上位性QTL分别是11对与14对,对表型变异的贡献率为0.04%～36.46%;这些QTL的表达呈现出时空性与特异性,即多数QTL仅在发育的一个特定时期表达,没有检测到能在整个灌浆期持续表达的QTL。在5A染色体的WMC74-Xgwm291-Xgwm410标记区间之间检测到多个控制各器官氮素含量的QTL,包括:QSnit-5A-1,Qknit-5A-2,Qknit-5A-3,Qpnit-5A-1;以及在3D染色体Xgwm341-Xgwm456标记区间检测到多个控制各器官氮素含量的QTL,包括Qlnit-3D-2,Qlnit-3D-3,Qlnit-3D-4;深入发掘这些热点区域,对于后期进行精细定位与图位克隆具有重要意义。4.两种水分条件下,花后不同发育阶段,共检测到氮素积累转运的32个加性QTL与23对上位性QTL,对表型变异的贡献率在5.39%～18.47%之间。其中检测到控制花后氮素积累量的5个加性QTL和2对上位性的QTL,对表型变异的贡献率分别为5.47%-10.96%和0.01%-6.31%;检测到控制花前氮素转运量的3个加性QTL和1对上位性QTL,对表型变异的贡献率分别为12.26%-17.81%和4.29%;检测到控制花前对籽粒氮积累贡献率的3个上位性QTL和3对上位性QTL,对表型变异的贡献率为9.64%-15.55%和0.47%-16.13%;检测到控制花后氮素转运量的2个加性QTL和3对上位性QTL对表型变异的贡献率分别为5.62%-10.35%和9.47%-18.79%;检测到控制花后氮素转运率的1个加性QTL和2对上位性QTL,对表型变异的贡献率分别为9.66%,和14.75%-24.38%;检测到控制花后对籽粒氮积累的贡献率的4个加性QTL和1对上位性QTL,对表型变异的贡献率分别为5.39%-15.55%和19.96%;检测到控制氮素籽粒生产效率的3个加性QTL和1对上位性QTL,对表型变异的贡献率分别为7.43%-16.15%和16.36%;检测到控制氮素干物质生产效率的11个加性QTL和10对上位性QTL,对表型变异的贡献率分别为5.75%-34.91%和0.82%-35.22%。在3D染色体上的Xgwm456-Xgdm8标记区间以及1B染色体的P3470.4-P4133标记区间检测到控制多个性状的主效QTL,且对表型具有较高的遗传贡献率,具有应用于MAS育种的前景。更多还原

【Abstract】 China is a large country consuming nitrogen fertilizers,but simultaneously,its using efficiency of nitrogen fertilizers is always lower than that of average level of the world.In the precondition that the total application of nitrogen for wheat is constantly added,the nitrogen use efficiency of wheat must be increased to maintain high yield of wheat.Screening genotype of high-efficiency of nitrogen is the main approach to improve using efficiency of nitrogen fertilizers and lessen environmental pollution.In this study,sixteen wheat cultivars were used as tested materials,the principal component analysis and correlation were conducted to find indexers which can be used to accurately evaluate using efficiency of nitrogen,then cluster was conducted with combination of the indexes and yield traits to screen cultivars with different using efficiency of nitrogen,and physiological and biochemistrical characteristics of the cultivars screened were analyzed for revealing genetic mechanism of accumulation and transportation of nitrogen;QTL was applied by accumulation and transportation of nitrogen of DH population for searching QTL distribution hot spots that can stable expression,which will present references for breeding for wheat cultivars with high using efficiency of nitrogen in the future.The main results of the study are as following: In this study,16 wheat cultivars were used as tested materials and total 14 nitrogen accumulation and translocation related traits,including quantity of pre-flowering nitrogen accumulation,quantity of post-flowering nitrogen transportation accumulation,quantity of leaf nitrogen content in mature period/ quantity of total nitrogen accumulation,quantity of rachilla nitrogen content in mature period/ quantity of total nitrogen accumulation,quantity of kernel nitrogen content in mature period/ quantity of total nitrogen accumulation and so on,were measured.After the indicators with larger coefficient of variation were selected,8 indexes related to using efficiency of nitrogen were identified by the principal component analysis and correlation,they were quantity of post-flowering nitrogen accumulation,quantity of pre-flowering nitrogen transportation,contribution rate of pre-flowering nitrogen transportation,quantity of post-flowering nitrogen transportation,rate of post-flowering nitrogen transportation,contribution rate of post-flowering nitrogen transportation,production efficiency of nitrogen in seeds,production efficiency of dry matters of nitrogen.The 16 wheat cultivars were divided into 3 categories according to cluster analysis by using the 8 indexes and the indexes relevant to yield.The first category has high nitrogen use efficiency,including Jinmai 54 and Jinmai 66,and the third category has loe nitrogen use efficiency,including Jinmai 73 and Taimai 269,while the second category is the intermediate type,including Jinmai 61,Tainong 18.Two varieties were chosen from each category,a total of six varieties,to mensurate activity of nitrogen accumulation and translocation related enzyme.The results showed that the activities of the following substences such as soluble protein,glutamine synthetase,glutamate synthetase,transglutaminase,and aspartate kinase had descending tendency after 7 days of flowering,while proteolytic enzyme showed ascending tendency after 7 days of flowering.The enzyme activities of Jinmai 54 and Tainong 66 were higher than those of Jinmai 73,Taimai 269,Jinmai 61,and Tainong 18 in grain filling stage.Correlation between enzyme activities and using efficiency of nitrogen indexes indicated that the activities of most of the crucial metabolic enzymes in flag leaves of the cultivars tested in the different stages of grain filling stage after flowering had significant or extremely significant genetic positive correlation with the 6 main indexes of using efficiency of nitrogen including production efficiency of nitrogen in seeds,production efficiency of dry matters of nitrogen,quantity of post-flowering nitrogen transportation,efficiency of post-flowering nitrogen transportation,contribution rate of post-flowering nitrogen accumulation in seeds,and had significant or extremely significant phenotype and genetic positive correlation with them but had significant or extremely significant phenotype and genetic negative correlation with the 2 main indexes of absorption and utilization of nitrogen including contribution rate of pre-flowering nitrogen accumulation in seeds,quantity of pre-flowering nitrogen transportation.Additionally,genetic correlational coefficiencies between the activities of most of the crucial metabolic enzymes and the main indexes of using efficiency of nitrogen were higher than phenotypic correlational coefficiencies,which suggested that the traits had excellent genetic correlation.which suggested that the expression of these 6 genes do involved in the accumulation and translocation of nitrogen in wheat.Under two water supply treatments,in different developmental stages after flowering,nitrogen content QTL of different developmental stages of leave,stem,rachilla,kernel and plant were detected by detection of unconditional and conditional QTL,additive QTLs controlling nitrogen content in leaves were respectively 10 and 7 with 5.18%~23.8% contribution rates to phenotypic variation,epistatic QTLs were 18 and 11 with 0.50%~43.76% contribution rates to phenotypic variation.Additive QTLs controlling nitrogen content in stems were respectively 12 and 14 with 6.72%~19.67% contribution rates to phenotypic variation,epistatic QTLs were 19 and 7 with 0.88%~33.63% contribution rates to phenotypic variation.Additive QTLs controlling nitrogen content in rachillae were respectively 5 and 4 with 8.65%~17.56% contribution rates to phenotypic variation,epistatic QTLs were 9 and 9 with 0.81%~38.72% contribution rates to phenotypic variation.Additive QTLs controlling nitrogen content in kernels were respectively 14 and 5 with 6.08%~27.44% contribution rates to phenotypic variation,epistatic QTLs were 9 and 5 with 0.40%~30.34% contribution rates to phenotypic variation.Additive QTLs controlling nitrogen content in plants were respectively 15 and 11 with 6.07% ~ 23.06% contribution rates to phenotypic variation,epistatic QTLs were 11 and 14 with 0.04% ~ 36.46% contribution rates to phenotypic variation.The expression of the QTLs showed specialty on space and time,that is to say,most of the QTLs merely expressed in a specific developmental stage and QTLs constantly expressing the whole grain filling stage were not detected.Many main QTLs controlling nitrogen accumulation quantity were detected in the marking regions of WMC74-Xgwm291-Xgwm410 in the chromosome 5A including Qlnit-3D-2,Qlnit-3D-3,Qlnit-3D-4.Deep probe of these hot spots is meaningful for accurate location and mapping clone in the late stage.Under two water supply treatments,in the different developmental stages after flowering,32 additive QTLs and 23 epistatic QTLs controlling nitrogen transportation with 5.39%~18.47% contribution rates to phenotypic variation were detected.Among them,5 additive QTLs and 2 epistatic QTLs controlling quantity of post-flowering nitrogen accumulation with 5.47%~10.96% and 0.01%~6.31% contribution rates,respectively,to phenotypic variation were detected;3 additive QTLs and 1 epistatic QTL controlling quantity of pre-flowering nitrogen transportation with 12.26% ~ 17.81% and 4.29% contribution rates,respectively,to phenotypic variation were detected;3 additive QTLs and 3 epistatic QTLs controlling rate of pre-flowering contribution of nitrogen accumulation in seed with 9.64%~15.55% and 0.47%~16.13% contribution rates,respectively,to phenotypic variation were detected;2 additive QTLs and 3 epistatic QTLs controlling quantity of post-flowering nitrogen transportation with 5.62%~10.35% and 9.47%~18.79% contribution rates,respectively,to phenotypic variation were detected;1 additive QTL and 2 epistatic QTLs controlling rate of post-flowering nitrogen transportation with 9.66% and 14.75%~24.38% contribution rates,respectively,to phenotypic variation were detected;4 additive QTLs and 1 epistatic QTL controlling rate of post-flowering contribution of nitrogen accumulation in seed with 5.39%~15.55% and 19.96% contribution rates,respectively,to phenotypic variation were detected;3 additive QTLs and 1 epistatic QTL controlling production efficiency of nitrogen in seeds with 7.43%~16.15% and 16.36% contribution rates,respectively,to phenotypic variation were detected;11 additive QTLs and 10 epistatic QTLs controlling production efficiency of dry matters of nitrogen with 5.75%~34.91% and 0.82%~35.22% contribution rates,respectively,to phenotypic variation were detected.Main effect QTLs controlling multi-traits were detected in the marking regions such as Xgwm456-Xgdm8 in the chromosome 3D and P3470.4-P4133 in the chromosome 1B.These QTLs had high contribution rates to phenotypic variation,which should have prospect in MAS breeding.更多还原