Starch is an insoluble glucose polymer that forms semi-crystalline granules. Insight into the relationships between the physicochemical properties of starch and its structure is limited. A deeper knowledge of these relationships is necessary for understanding the starch properties of mutant lines. Starch synthase (SS) and starch branching enzyme (BE) play central roles in the biosynthesis of starch. To explore the relationships between the physicochemical properties of starch, morphology of starch granules, starch crystallinity, and the function of SS and BE isozymes expressed in rice endosperm, we analyzed these traits in starches isolated from the double mutant lines [ ss1/be1 and ss1^L/be2b ( ss1^L means leaky ss1 mutant)]. Deficiency of SSI and BEI lead to an increase and decrease in gelatinization temperature, respectively. A significant increase in gelatinization temperature results from deficiency of BEIIb, as has been shown in previous studies in rice and maize, and this phenomenon is pronounced in the reduction of SSI activity. Deficient BEIIb expression also had a significant impact on the morphology of starch granules in the endosperm and changed the A-type diffraction pattern of the granules to a B-type pattern, while deficiency of SSI and/or BEI did not affect these traits.