棉花光合关键基因RCA的eQTL定位及优异等位变异挖掘

Rubisco activase, one of the key photosynthetic enzyme, that regulate the activity of Rubisco in plant. Numerous studies have indicated that endogenous levels of RCA could serve as an important determinant of photosynthetic efficiency, plant productivity and stress response. However, the molecular mechanism of transcriptional regulation about RCA genes has thus far remained unknown. In the previous work of this project, we have cloned GhRCAα and GhRCAβ, and analyzed the tissue expression of two RCA genes. It was found that GhRCAα and GhRCAβ in cotton were highly expressed in leaves, but that their expression pattern were significantly different. Compared with GhRCAα, GhRCAβ was expressed at higher mRNA and protein levels. On this basis, this study will detect expression quantitative trait loci (eQTL) for RCA genes and quantitative trait loci (QTL) for yield related traits using recombinant inbred lines population and further investigate the relationship between the effect of RCA genes expression on cotton photosynthesis and productivity. Simultaneously, through the association analysis on the key eQTL interval in natural population, we will detect the excellent allelic variation and optimal haplotype, and develop functional markers closely linked with RCA genes expression. These results could provide a new train of thought for increasing overall cotton photosynthesis and productivity by regulating expression of RCA genes through marker-assisted breeding.

Rubisco活化酶（RCA）是植物光合作用过程中的关键酶，具有调节Rubisco活性的功能。已有大量研究表明RCA基因对于植物的光合效率、产量形成和逆境响应具有重要作用。然而，人们对调控RCA基因表达的分子遗传机制还知之甚少。本项目前期克隆了棉花的两个RCA基因并分析了其组织表达规律，结果发现GhRCAα和GhRCAβ在叶片中表达量最高但具有不同的表达模式。相比GhRCAα，GhRCAβ在mRNA水平和蛋白水平具有较高的表达。本项目在此基础上利用一套棉花重组自交系群体（RIL）全基因组鉴定调控RCA基因表达的eQTL，并定位产量QTL，进一步分析两个RCA基因与产量之间的关系；同时，在自然群体中鉴定关键eQTL区段的优异等位变异和最优单倍型，开发与棉花RCA基因表达水平相关的分子标记或功能标记。研究结果可为育种上通过分子标记手段调控RCA基因表达，实现棉花光合能力和产量的提高提供新思路。

Rubisco活化酶（RCA）是催化植物中Rubisco活化的关键酶。通过调控RCA基因的表达增加Rubisco的活性，对于改善植物的光合效率，从而提高植物的产量具有重要意义。本项目克隆了棉花RCA基因GhRCAβ2，并对GhRCAβ2表达调控的遗传机制进行了研究。研究结果表明：（1）Western-blot结果显示GhRCAβ2重组蛋白可与RCA抗体发生免疫反应，表明GhRCAβ2的确是编码RCA蛋白的基因。（2）亚细胞定位结果表明GhRCAβ2是一个定位于叶绿体的蛋白，这与其主要参与光合作用的功能一致。（3）组织表达分析发现GhRCAβ2在光合作用进行的主要位置叶片中表达量最高，且在黄绿叶突变体中的表达显著低于对照；进一步诱导表达分析表明GhRCAβ2的表达受低钾和低磷胁迫诱导。（4）利用一套自然群体对GhRCAβ2表达与光合性状进行相关分析发现，GhRCAβ2表达与叶绿素荧光参数qP呈显著正相关，与NPQ呈显著负相关；（5）进一步eQTL定位分析发现，共鉴定到23个与GhRCAβ2表达显著关联的SNPs，但是未鉴定到与叶绿素荧光参数共定位的SNPs。（6）对GhRCAβ2启动子进行序列分析发现，GhRCAβ2启动子区除具有TATA-box和CAAT-box等基本顺式作用元件外，还含有多个响应于光、逆境胁迫和植物激素等顺式作用元件。本研究结果有助于深入了解GhRCAβ2表达调控的分子机制，并为棉花产量的提高及高光效育种提供理论依据。