高直链淀粉水稻胚乳中空状和细长状淀粉粒的发育和结构特性

High-amylose cereal kernels contain a high level of resistant starch and have a great benefit on human health. They are an important source of functional foods and have potential market demand and application prospect. Studies at home and abroad have shown that the proportion of the internal hollow and elongated starch granules in cereal endosperm starches increases with the significant increase in amylose content. However, the development and structural properties of these starch granules are not reported..A high-amylose transgenic rice line (TRS) has been developed by antisense RNA inhibition of the starch branching enzymes. The high proportion of the internal hollow and elongated starch granules in TRS endosperm starches has been observed in our previous research. In this project, the formation, developmental stage and distribution of internal hollow and elongated starch granules in TRS endosperm will be investigated with light microscopy, electron microscopy and confocal laser scanning microscopy during endosperm development. The relationship between starch synthesis key enzymes and the development of internal hollow and elongated starch granules will be studied with histochemical, immunocytochemical and proteomics techniques during TRS endosperm development. Native internal hollow and elongated starch granules will be isolated from TRS mature kernels, and their crystalline and structural properties will be investigated by a combination of morphological and spectroscopic analyses and physicochemical determination. The research results will be very useful for breeding of high-amylose cereal crops and various applications of high-amylose cereal starches in the food and nonfood industries. The results can also add to our understanding of development and structural properties of plant starch granule.

高直链淀粉作物籽粒富含抗性淀粉，具有促进健康的功能，可作为功能性食品有着潜在的市场需求和应用前景。国内外的研究都表明，直链淀粉含量显著提高后，禾谷类作物胚乳中空状和细长状淀粉粒的含量随之增加，而这些淀粉粒的发育和结构特性尚未澄清。.课题组在前期工作中，观察到基因工程培育出来的高直链淀粉水稻TRS胚乳富含中空状和细长状淀粉粒。本项目拟利用光镜、电镜和激光共聚焦显微镜等技术，在TRS胚乳发育过程中，研究并探明中空状和细长状淀粉粒的形成过程、发生时期和在胚乳组织中的空间分布。利用组织化学、免疫细胞化学和蛋白质组学等手段，研究并阐明淀粉合成关键酶与中空状和细长状淀粉粒发育的关系。从TRS成熟籽粒中分离纯化中空状和细长状淀粉粒，进行波谱分析和理化测定，研究并查明它们的晶体和结构特性。研究结果可以为高直链淀粉作物的选育、高直链作物淀粉的开发和应用提供基础理论，并充实植物淀粉粒的发育和结构等学科知识。

高直链淀粉作物籽粒富含抗性淀粉，具有促进健康的功能，可作为功能性食品有着潜在的市场需求和应用前景。然而直链淀粉含量显著提高后，胚乳细胞中出现多聚体、细长状、中空状等不同形态淀粉粒(简称为异形淀粉)，并且细长状和中空状淀粉的含量随着直链淀粉含量的升高而增加。而这些异形淀粉的发育和结构特性尚未澄清。.本项目主要以富含异形淀粉的高直链淀粉水稻TRS为材料，建立了淀粉发育和特性的原位分析技术以及异形淀粉的分离纯化方法，观察了TRS异形淀粉的形成过程和在胚乳组织中的时空积累，研究了淀粉合成关键酶与异形淀粉发育的关系，测定了异形淀粉的分子结构、晶体结构和功能特性。.获得的主要结果如下：(1) 原位分析技术的建立为研究异形淀粉的发育和特性提供了保障，异形淀粉分离纯化方法的建立为测定异形淀粉结构和功能特性奠定了基础。(2) TRS胚乳有多角形、多聚体、细长状和中空状等异形淀粉。多角形淀粉为复粒淀粉的亚颗粒，分布于胚乳中央，在花后5 d之前形成；多聚体淀粉由复粒淀粉外围亚颗粒彼此融合形成，分布于中央胚乳和亚糊粉层之间，在花后5-10 d形成；细长状淀粉由线状排列的复粒淀粉亚颗粒融合形成，分布于亚糊粉层内侧的胚乳细胞，在花后5-16 d形成；中空状淀粉是无定形淀粉在淀粉体内膜处沉积形成，分布于亚糊粉层，在花后7-16 d形成。(3) 伴随胚乳发育，从籽粒内部到外部淀粉分支酶的表达抑制越来越显著，支链淀粉分支度逐渐降低和侧链逐渐变长，淀粉从多角形逐渐形成多聚体、细长状和中空状淀粉。(4) 从多角形到多聚体、细长状和中空状淀粉，直链淀粉含量逐渐升高，支链淀粉分支度降低但侧链变长，淀粉晶体由A-型经C-型到无定形，淀粉抗糊化能力逐渐增强。细长状和中空状淀粉表现为明显的双相酶水解曲线，而多角形和多聚体淀粉有单相酶水解曲线，且多角形淀粉抗酶水解最弱、细长状淀粉最强。.该研究为高直链淀粉作物的选育、高直链作物淀粉的开发和应用提供基础理论，并充实植物淀粉的发育和结构等学科知识。