玉米Rubisco活化酶基因eQTL的遗传解析及育种利用研究

Rubisco activase regulates the activity of Rubisco, and is a key enzyme of photosynthesis in plant. Numerous studies have shown that RCA gene plays an important role in plant light use efficiency and yield. Therefore, RCA gene has become a potential target for improving plant performance. However, to date, little information about the genetic architecure that regulates RCA gene is available. In our previous study, we cloned two RCA genes from maize. Correlation analysis in 123 maize inbred lines showed that the expression level of these two genes was correlated positively with grain yield. The correlation with grain yield for the expression of one of the two genes was even larger than that for three C4-specific photosynthesis high-efficiency genes that are widely studied in maize. In total, the two RCA genes explained approximately 24% of the grain yield variation in the 123 inbred lines. Our preliminary expression quantitative trait loci (eQTL) analysis showed that RCA genes were controlled by both Trans- and Cis-eQTLs, and some eQTLs were co-localized with the QTL for grain yield. Based on the above findings, we will carry out the following studies in this research project: (1) Identify the key genomic regions that regulate the expression level of RCA genes and grain yield, through both linkage and association analysis, in a maize recombinant inbred line (RIL) population and a maize association mapping panel, and dissect the genetic base for the observed phenomenon that RCA gene expression is closely correlated with grain yield. (2) Screen for elite alleles for RCA genes and the key genomic regions as were identified above, using candidate association approach. (3) Confirm the functional properties of the candidate elite RCA gene promoter alleles through genetic transformation way. (4) Develop new maize materials through marker-assisted selection methods, which pyramid the identified elite alleles from different loci to inbred lines of interest to produce superior genotypes, and evaluate the breeding value of these elite alleles. The results gained from this research project will not only enhance the understanding of genetic mechanism that underlies photosynthesis and grain yield, but also provide a new idea to develop maize hybrids with high yield in breeding practice.

Rubisco是光合作用限速酶，但必须在其活化酶（RCA）催化下才具有活性。RCA对植物光能利用和产量建成具有重要作用，已成为作物高产育种潜在靶点。我们前期克隆出2个玉米RCA基因；在123份玉米自交系中，其表达对产量贡献率高达24%，其中1个基因与产量间相关系数高于C4途径3个重要高光效基因；基因定位初步结果显示，2个RCA基因均具有与产量QTL共位的eQTL。本项目拟在此基础上，通过连锁分析和全基因组关联分析，鉴定调控玉米RCA基因表达量和产量性状的关键基因组区段，剖析RCA基因参与产量建成的遗传基础；通过目标区间定点关联分析，筛选关键基因组区段的优异等位变异；利用基因瞬时表达方法，验证RCA基因启动子的优异等位变异；并利用标记选择方法，将优异等位变异导入需要改良的自交系，评价其育种价值。研究结果对于解析玉米光合作用和产量建成的遗传基础，培育高产品种具有重要理论意义和应用价值。

Rubisco活化酶（RCA）对植物光合作用和产量建成具有重要作用。本项目采用连锁分析和全基因组关联分析方法，定位调控玉米RCA基因表达量和产量性状的关键基因组座位，挖掘关键座位的优异等位变异，利用转基因方法验证优异等位变异，并开展优异等位变异的分子标记辅助育种工作。研究进展如下：①以242份玉米重组自交系为材料，在连续两年的大田种植条件下，检测到5个控制RCA基因ZmRCAβ表达的eQTL和4个产量QTL。以具有广泛遗传变异的235份玉米自交系为材料，在连续两年的大田种植条件下，检测到15个控制ZmRCAβ基因表达的eQTL和2个产量QTL。两项分析共同检测到一个在不同环境中稳定表达的eQTL。在所有检测到的位点中，该eQTL的遗传效应最大，对表型变异的最高贡献率达29.7%。②因具有最大遗传效应的eQTL与ZmRCAβ基因物理位置重叠，我们推测，ZmRCAβ基因表达受其自身DNA序列变异调控。以182份玉米自交系为研究材料，对ZmRCAβ基因启动子区、编码区和非编码区开展DNA测序和候选区段关联分析，发现基因启动子区存在2个与ZmRCAβ基因表达显著关联的多态性位点，它们分别为一个2-bp插入和一个14-bp的插入。依据这两个插入将ZmRCAβ基因启动子划分成3种单倍型，其中携带ZmHap1单倍型的自交系，其RCA基因表达量和产量高于携带其它单倍型的自交系。在水稻愈伤组织、玉米和水稻原生质体、烟草叶片中的转基因实验进一步证实，ZmHap1型启动子活性强于其它类型启动子。③设计了2个基于PCR的分子标记，并通过分子标记辅助选择方法获得3份了导入优异等位变异的玉米育种中间材料。上述研究结果对于解析玉米光合作用形成的遗传基础和培育高产玉米新品种具有重要理论和应用价值。本项目执行期间共发表研究论文7篇，其中SCI论文5篇。培养硕士研究生8名。