High-amylose cereal starch has a great benefit on human health through its resistant starch content. In this paper, starches were isolated from mature grains of high-amylose transgenic rice line (TRS) and its wild-type rice cultivar Te-qing (TQ) and digested in vitro and in vivo. The structural changes of digestive starch residues were characterized using DSC, XRD, (13)C CP/MAS NMR, and ATR-FTIR. TQ starch was very susceptible to digestion; its residues following in vitro and in vivo digestion showed similar structural characteristics with TQ control starch, which suggested that both amorphous and crystalline structures were simultaneously digested. Both amorphous and the long-range order structures were also simultaneously hydrolyzed in TRS starch, but the short-range order (double helix) structure in the external region of TRS starch granule increased with increasing digestion time. The A-type polymorph of TRS C-type starch was hydrolyzed more rapidly than the B-type polymorph. These results suggested that B-type crystallinity and short-range order structure in the external region of starch granule made TRS starch resistant to digestion.