普通小麦阿魏酰阿拉伯木聚糖含量主效QTL QFax.xjau-3AS精细定位和物理图谱构建

Arabinoxylans (AX) belong to an important group of non-starch polysaccharides in wheat grain, and they significantly affect the properties of dough and end-use products of wheat. Arabinoxylan feruloyl transferase (AFT), a critical enzyme in the arabinoxylans biosynthetic pathway, demonstrated high association with feruloyl arabinoxylan (FAX) content in wheat grain. FAX content in grain is an important factor in determining the structure of AX. Previously, we identified a major QTL QFax.xjau-3AS for FAX content in Gaocheng 8901/Zhoumai 16 RIL (recombinant inbred line) population using the wheat 90K SNP assay. In the present study, the F2, F2:3, F3:4 and F4:5 secondary populations from the cross of GZ5818×GZ5616 will be used for genetic analysis based on the flanking markers of QFax.xjau-3AS and the linked markers of other QTLs (GZ5818 has an allele at QFax.xjau-3AS locus for higher FAX content whereas GZ5616 has an allele for lower FAX content; two parents showed only allelic variations at QFax.xjau-3A locus). The BSR-Seq technique and genomic sequence and physical map information of chromosome 3A will be employed to develop molecular markers tightly linked (with genetic distances less than 0.1 cM) or co-segregated with QFax.xjau-3AS for a high-resolution genetic mapping. In addition, a physical map covering QFax.xjau-3AS will be constructed and candidate genes will be detected. This study will provide useful information for marker-assisted selection in wheat breeding and map-based cloning of QFax.xjau-3AS.

阿拉伯木聚糖（AX）是小麦中重要的膳食纤维组分，对面制品加工品质和营养品质有重要影响。阿魏酰阿拉伯木聚糖（FAX）含量是决定小麦AX质量的关键因素。项目申请人前期利用90K SNP芯片在藁城8901/周麦16重组自交系群体中发现一个FAX含量的主效QTL QFax.xjau-3AS，本项目以该QTL侧翼标记为参考，用携带该位点高FAX含量等位基因的纯合株系GZ5818和低FAX含量等位基因的纯合株系GZ5616组合（双亲间只在该主效QTL位点有差异）的F2、F2:3、F3:4和F4:5群体为研究材料，通过3个世代的混池RNA测序（BSR-Seq），并结合3A染色体基因组序列信息，发掘与该位点紧密连锁（连锁距离0.1 cM以内）或共分离的分子标记，构建覆盖该位点的高精度遗传图谱和物理图谱，精细定位QFax.xjau-3AS，并发掘其候选基因，为分子标记辅助育种及该基因的图位克隆奠定基础。

在藁城8901/周麦16群体中共检测到5个 QTL，QFax.xjau-2AS、QFax.xjau-2BS、QFax.xjau-2DL、QFax.xjau-3AS和QFax.xjau-6A，分别位于2AS、2BS、2DL、3AS和6AS染色体上。QFax.xjau-2AS、QFax.xjau-2BS、QFax.xjau-2DL、QFax.xjau-3AS和QFax.xjau-6AS在不同环境下能分别解释6.2-18.2%的表型变异。基于全基因组关联分析，检测到110个分布于21个染色体区域（-log10 (P) ≥3.0）与FAX含量显著相关的SNP，单个SNP 可以解释6.8-21.2%的表型变异。在显著相关的SNP 位点区域内发现5个候选基因。利用同源克隆的方法克隆了普通小麦3A、3B、3D染色体上AFT基因的全长编码序列，分别命名为TaBahd-A1、TaBahd-B1和TaBahd-D1。在 TaBahd-A1，TaBahd-B1和TaBahd-D1基因点均发现2-3个等位变异，分别据等位变异类型开发了一对各自显性互补的功能标记AFTA2和AFTB2，AFTB7和AFTC8，AFT 97和AFT 85，具有TaBahd--A1a、TaBahd--B1a 和 TaBahd--D1a 的材料与高 FAX含量相关，而具有TaBahd--A1b, TaBahd--B1b and TaBahd--D1b 型的材料与低FAX含量相关。用基于TaBahd-A1、TaBahd-B1和TaBahd-D1 三个基因位点开发的两对显性互补的标记相结合，进一步对冬小麦材料进行检测，显著提高了标记的选择效率，用TaBahd-A1a / TaBahd-B1a / TaBahd-D1a更能有效选择高FAX含量的材料。采用半定量RT-PCR技术对TaBahd-A1、TaBahd-B1和TaBahd-D1基因的时空表达模式进行了分析，TaBahd-A1、TaBahd-B1和TaBahd-D1基因在籽粒中特异表达，均在开花后21表达水平最高，随着籽粒成熟表达水平迅速降低。3个染色体TaBahd基因同时发生基因编辑的植株籽粒FAX含量为3.57×10-5 mol/L，低于受体Fielder品种（4.07×10-5 mol/L）。初步验证了TaBahd基因可能与胚乳细胞壁FAX含量存在相关性。