Effects Of Grain Quality By Inhibition The Expression Of Different Starch Branching Enzymes In Rice

Rice (Oryza sativa L.) is one of the people’s main food crops in the world. The rice endosperm is rich of starch, and starch is divided into amylose and amylopectin. The proportion of amylose and amylopectin and amylopectin structure jointly determine the quality and features of rice grains. Starch branching enzyme (SBE) is one of the key enzymes involved in starch synthesis, which is the only branch introduced in the process of starch synthesis enzymes. Rice SBE has three kinds of isforms, named as SBEI (SBEI), SBEIIa (SBE4) and SBEIIb (SBE3). Existing research shows that SBEI and SBE3 mainly involved in the synthesis of branched chain starch in the endosperm, and SBEI transfers a long branched chain in rice during whole growth period, while SBE3 mainly transfers short branched chain during the process of the branched chain starch synthesis in rice endosperm. SBE4 mainly involved in the vegetative organs such as blade branched chain starch synthesis, it also have a small amount of expression in the endosperm, but the mechanism works on physicochemical properties and starch structure of rice is still unclear.In the previous works of, the SBE4 RNA interference construct, and the antisense structure for both SBEI and SBE4, or both SBE3 and SBE4 genes was introduced into the japonica rice variety Yandao 8 mediated by agrobacterium, respectively. Based on these transgenic plants with different background of SBE inhabitation, this study carried out the molecular identification and expression analysis of the homozygous lines containing the target gene.And different background homozygous transgenic lines were selected to compare the physical and chemical quality and starch fine structure. The main research results summarized as follows:1. By using hygromycin resistance test of leaf and PCR identification method, the results showed that the target gene has been integrated into the transgenic rice genome; The data from Real-time PCR analyses showed the endogenous SBE genes’expression in developing seeds of transgenic rice was significantly lower than that of control. Compared with those in parent, the expression of SBE1 and SBE3 genes dropped significantly. And SBE4 gene expression also decreased, but the change level was lower than the previous two genes, this might be due to the low expression level of endogenous SBE4 gene in rice grains.2. The results from field trial implied that the transgene(s) had no obvious effect on the agronomic traits such as plant height, tiller number, spike length and spike grain number, but had a significant impact on grain appearance and thousand-seed weight. The chalk white degree oftransgenic grains increased, grain weight and thickness significantly decreased. The data of general quality analyses showed that the apparent amylose content and the resistance starch content of transgenic rice were obviously increased, especially in transgenic rice with inhibiting SBE3 and SBE4. The results from the starch viscosity and thermodynamic properties indicated that the pasting temperature of transgenic rice increased, the viscosity descreased, implied that the transgenic rice grains with different background of genetically modified SBEs present enhanced ability to resistance of pasting.3. By using scanning electron microscopy (SEM) to invectigate the cross-section of rice grains, the results showed that the grain internal gap increased, starch granules easily cracks, the surface was porous, and the starch granules was arranged loosely. It was implied that there was largest change of the starch granules in transgenic plants containing the anti-SBE3/4 structure.4. Use high temperature gel exclusion chromatography (GPC) analysis, the data showed that the transgenic rice starch contains more amylose and long branch amylopectin chains, but less short branch amylopectin chainss.