小麦淀粉合成酶基因TaSSIV新等位变异挖掘及其在淀粉合成中的遗传作用

Starch has been playing important roles in human life and food consumption, it is synthesized by several dozen starch synthase enzymes, SSIV is one of the isoforms, its genetic function on synthesis of amylose /amylopectin /resistant starch in wheat is not so clear due to lack of respective mutant lines..We have identified five novel alleles of gene TaSSIV in D sub-genome, and the mutations were in functional and conserved domains, which resulted in the changes of amino acid, and led to significant down regulation of its transcript in leaves and seeds, starch granules in chloroplasts were reduced remarkably. By using TILLING platform and the EMS mutagenized population, this proposal is going to identify novel alleles of TaSSIV in A and B sub-genome, to pyramid novel alleles of the three sub-genomes, and to breed double /triple mutants of TaSSIV. By integrated application of RT-PCR, Western blot, electron microscope and other techniques, expression pattern of TaSSIV and other starch synthesis related genes will be analyzed to investigate their effects on amylose /amylopectin /resistant starch in wheat. The results will explicate the genetic effects of gene TaSSIV on amylose /amylopectin /resistant starch, illustrate its biological mechanism in storage starch synthesis, and reveal the dosage and pleiotropic effects among homologous and other genes. The study will provide experimental data for comprehensively interpreting function of TaSSIV in wheat, and promote research on homologous gene evolution.

淀粉是由多种淀粉合成关键酶类共同催化调控合成的，SSIV酶即是其中之一。目前由于缺乏有效的SSIV同源基因突变材料，对该基因在小麦淀粉合成路径中的遗传与互作机制、直链/支链/抗性淀粉含量的贡献率尚不十分明确，亟待深入研究。.本团队前期鉴定获得了小麦TaSSIV基因在D基因组上的5个位于功能域的新等位变异，突变导致其在叶片和籽粒中的表达显著降低，叶绿体中淀粉粒数量显著减少。本项申请拟在此基础上进一步挖掘TaSSIV基因在A、B基因组上的新等位变异，构建双/三突变体，对比分析单/双/三突变体籽粒中TaSSIV基因表达及其对其他淀粉合成关键基因的影响，研究淀粉粒特性和直链/支链/抗性淀粉变异，明确该基因在小麦籽粒淀粉合成中的遗传作用，阐明其在淀粉合成中的作用机制，揭示同源基因间及与其他关键基因的补偿/互作效应，为全面深入解析小麦SSIV基因功能提供实验数据，促进小麦同源基因互作机制的深入研究。

SSIV酶是淀粉合成关键酶类之一，共同催化调控淀粉合成。本项目研究团队以小麦京411突变体库为材料，采用定向诱导基因组局部突变技术（TILLING）挖掘到小麦淀粉合成酶基因TaSSIV在A和B两个染色体组上的5个功能性等位变异。与前期获得的TaSSIV-D终止突变聚合后，创制了双/三突变体和基因编辑株系，以单、双、三突变体为材料，分析了TaSSIV基因在不同突变体中的表达情况和对叶片、籽粒淀粉粒合成的影响。研究明确了TaSSIV基因的遗传作用和生物学功能，在叶片和籽粒中均有表达，但叶片中表达水平较高；该基因突变体后显著影响叶片中淀粉粒和籽粒淀粉的合成，TaSSIV-D的影响大于TaSSIV-A、B。明确了TaSSIV三个同源基因间的互作模式，TaSSIV部分同源基因表达模式在小麦叶片淀粉粒合成中存在反馈调节。TaSSIV-B、D对TaSSIV-A表达有极显著负反馈调节作用，且TaSSIV-B、D对TaSSIV-A的表达具有累加效应；TaSSIV-A、D对TaSSIV-B的表达有极显著负反馈调节作用；TaSSIV-A、B对TaSSIV-D有极显著正反馈调节作用。本研究明确了TaSSIV基因在小麦叶片和籽粒淀粉合成中的作用机制和遗传效应，丰富了小麦淀粉合成遗传网络，并促进多倍体小麦部分同源基因互作机制研究；创制了淀粉突变新材料，为小麦淀粉品质改良奠定了材料基础。