大麦穗轴节数主控基因HvNRN的精细定位、克隆和功能分析

Grain Number per Spike is one of the main components of barley yield, and increasing Number of Rachis Node (NRN) is an effective way to increase spikelet as well as Grain Number per Spike for barley. However, few studies have been conducted on genetics and molecular mechanisms involved in the development of barley NRN so far. In our previous study, qNRN, the main QTL for NRN of barley, was initially mapped on the chromosome 2H of barley based on the DH population derived from Nongai No.1 (with less NRN) and Nongdamai 012 (with more NRN). The current study will focus on fine mapping and map-based cloning of qNRN using the secondary population derived from the two parents and the DH population. The relationships between the candidate gene HvNRN and NRN of barley as well as the mechanisms involved in it will also be studied and discussed. Furthermore, association study will be conducted to identify the beneficial allele of HvNRN. This study will not only lay the foundation for illustrating the molecular mechanisms involved in the development of NRN of barley, but also provide valuable information for the application of the causal gene in genetic improvement of grain number per spike of barley and breeding of elite barley and wheat varieties with high yield potential through modern strategies.

穗粒数是大麦产量最重要的构成因子之一，增加穗轴节（小穗着生部位）数是提高每穗小穗数进而提高穗粒数的有效途径，但是目前对大麦穗轴节数形成的遗传学和分子生物学研究还比较少见。在前期研究中，我们以穗轴节数有显著差异的2个亲本（农矮1号和农大麦012）及其衍生的DH群体对该性状进行了初步定位，在2H上检测到一个主效QTL qNRN。本项目拟在此基础上，充分利用大麦基因组序列信息，结合高通量测序、生物信息学等方法，对该主效QTL进行精细定位，并对其候选基因HvNRN进行克隆和转基因功能验证。进一步利用关联分析筛选该基因的优异等位变异，并通过细胞组织学等手段初步探讨该基因参与大麦穗轴节数形成的途径。本项目将为解析大麦穗轴节数形成的机理奠定基础，同时为大麦及小麦等作物的穗粒数及产量性状的遗传改良提供优异基因资源。

穗粒数是大麦产量最重要的构成因子之一，增加穗轴节数是提高小穗数进而提高穗粒数的有效途径。本研究以1815D（原农矮1号，主穗穗轴节数16.4）和红90-9（原农大麦012，主穗穗轴节数30.7）及其衍生后代分离群体为研究对象，通过正向遗传学的方法对调控大麦穗轴节数QTL进行了定位，对其中的主效QTL qSRN1进行了图位克隆，并通过CRISPR/Cas9的策略对候选基因进行了功能验证。结果发现，位于2H染色体上的水稻LAX2同源基因E3泛素蛋白连接酶（HORVU.MOREX.r3.2HG0164020）为大麦穗轴节数调控基因，将其命名为SRN1。该基因具有转录激活活性，编码蛋白位于细胞核内，可以通过与HvLAX2互作调控下游基因的表达。进一步分析表明，SRN1不仅可以调控大麦穗轴节数，还可以调控粒长。SRN1编码区序列在1815D和红90-9间无差异，但是启动子区包含4个SNP和4个Indel。根据上述变异，可将392份大麦种质分为9种单倍型。关联分析表明，其中3个SNP位点（SNP-1884，SNP-1748和SNP-1699）的等位变异与穗轴节数相关，多穗轴节数和少穗节数等位基因分别来源于红90-9和1815D，分别命名为SRN1-90和SRN1-18。这3个SNP位点中，SNP-1884和SNP-1748为功能位点，与该基因在不同种质中的表达丰度有关。对SNP-1884和SNP-1748在中国和全球大麦品种中的分布进行分析发现，SRN1-90的频率显著高于SRN1-18，野生大麦中却相反。此外，SRN1-90主要分布在欧洲和中国，而SRN1-18主要分布在亚洲西部和澳大利亚。本研究为大麦、小麦等重要作物穗粒数及产量性状的分子遗传改良提供了优异基因资源。