Effects Of Expression Inhibition Of Starch Branching Enzyme On In Situ Gelatinization And Degradation Of Starch In Brown Rice

Cereal endosperm is rich in starch,which is not only the main source of carbohydrate intake for people,but also provides the main energy for grain germination and seedling growth.The expression inhibition of starch branching enzyme(SBE)can increase resistant starch in endosperm,which has many health benefits for humans.Therefore,many cereal crop lines with expression inhibition of SBE have been cultivated.The effects of expression inhibition of SBE have been widely investigated on structure and properties of isolated starch from endosperm.However,the effects of expression inhibition of SBE on in situ gelatinization and degradation of starch in brown rice are not clear.In this study,three convention rice cultivars with different amylose contents including Indica rice Te-qing,Japonica rice Wu-xiang 9915,and waxy rice Guang-ling-xiang-nuo,and their transgenic rice lines with inhibition of SBEI/IIb were chosen,and their mature kernels were used as experimental materials.The changes of morphology and physicochemical properties of starch were investigated during kernel cooking process andseedling growth.The objective of this study was to reveal the effects of expression inhibition ofSBE on in situ gelatinization and degradation of starch in brown rice,and clarify the reason of the effects.This study could provide important reference for the cultivation and utilization of crops with high resistant starch,and enrich the knowledge of starch structure and properties.The main results were shown as following.1.Observation on in situ gelatinization and degradation of starch in brown rice usingresin section method of whole kernel.The morphology changes of endosperm starch during rice kernel cooking process and seedling growth were observed using our established resin section of cereal whole kernel.The results showed that the cooked kernel and germinated seed could be sectioned into whole section of kernel with 2 μm thickness using resin section method.The section of cooked kernel stained with iodine could clearly show that the kernel was gelatinized from the outer region to the inner region of endosperm.During cooking process,the volume of starch granules increased,the amylose was leached,and starch granules gradually lost their granule morphology and adhered together.The section of germinated seed showed that endosperm starch was gradually degraded from the proximal to distal region of embryo and from the outer to inner in endosperm.2.The effects of expression inhibition of SBEI/IIb on in situ gelatinization of starch in brown rice.After brown rice kernels were cooked in a boiling water bath for different time,the cooking quality of kernel was investigated,the gelatinization process of starch was observed,and structural properties of gelatinized starch were determined.The results showed that convention rice kernels were completely gelatinized after 30 min of cooking,but rice kernels with expression inhibition of SBE were not gelatinized completely even after 60 min of cooking.The volume,wet weight and leached material of cooked kernel increased with increasing cooking time.The increase was larger in convention rice kernel than in rice kernel with expression inhibition of SBE.The kernel was gradually gelatinized from the outer to the inner of endosperm for convention rice.However,for rice kernel with expression inhibition of SBE,the outer region of endosperm was gelatinized firstly but was not gelatinized completely,and the inner region of endosperm was gelatinized completely.The homogeneous and polygonal compound starch in convention rice kernel was easily gelatinized and adhered together.The outer region of endosperm with expression inhibition of SBE had aggregate,elongated and hollow starch,which had high resistance to gelatinization and did not adhered together.The molecular weight distribution did not show significant changes during kernel cooking process.The A-type starch was easily gelatinized into amorphous starch in convention rice,but the crystalline structure in C-type starch of rice kernel with expression inhibition of SBE was not wholly disrupted during cooking process,leading to that the cooked kernels had high resistance to digestion.The above results indicated that expression inhibition of SBE changed the morphology and crystalline structure of rice starch,which had high resistance to gelatinization and increased the resistant starch of cooked kernels.3.The effects of expression inhibition of SBEI/IIb on in situ degradation of starch in brown rice.The seeds were germinated and grew in the dark only in ddH2O.The relationship between endosperm starch degradation and seedling growth was investigated,the degradation process of starch in endosperm was observed,and the structure and properties of residual starch in germinated kernel were determined during seedling growth.The results showed that there was a significantly positive relationship between starch degradation and seedling growth.The expression inhibition of SBE decreased the hydrolysis of starch in rice,which restrained the seedling growth.During seedling growth,starch was gradually degraded from the proximal to the distal region of embryo and from the outer to the inner in endosperm.The endosperm starch could be degraded completely in convention rice kernel.The aggregated,elongated and hollow starch in rice kernel with expression inhibition of SBE had high resistance to enzyme hydrolysis,leading to that the starch in the outer region of endosperm could not be degraded completely.In convention rice,the starch components including amylose and amylopectin were simultaneously degraded,and starch crystalline structure did not change during seedling growth.However,in rice kernels with expression inhibition of SBE,the amylose was degraded faster than amylopectin,the amylopectin long branch-chains had high resistance to enzyme hydrolysis,and the A-type crystallinity in C-type starch was degraded faster than B-type crystallinity.The above results indicated that the long branch-chains of amylopectin and B-type crystallinity in rice endosperm with expression inhibition of SBE resisted in situ degradation and inhibited plant growth during seedling development.