¡¾ABSTRACT¡¿ Proteomic approaches were applied in four grain developmental stages of the Chinese Yunong 201 and its (EMS) mutant line Yunong 3114. 2-DE and tandem MALDI-TOF/TOF-MS analyzed proteome characteristics during middle and late grain development of the Chinese Yunong 201 and its EMS mutant line Yunong 3114 with larger grain sizes. We identified 130 differentially accumulated spots representing 88 unique , and four main expression patterns displayed a dynamic description of middle and late grain . Those identified species participated in eight biochemical processes: stress/defense, , /assembly/, storage , energy production and transportation, , transcription/, . Comparative proteomic characterization demonstrated 12 spots that co-accumulated in the two cultivars with different expression patterns, and six cultivar-specific spots including serpin, small heat shock ,β-amylase, α-amylase inhibitor, dimeric 1717-amylase inhibitor precursor, and cold regulated . These cultivar-specific spots possibly resulted in differential yield-related traits of the two cultivars. Our results provide valuable information for dissection of molecular and genetics basis of yield-related traits in and the proteomic characterization in this study could also provide insights in the biology of middle and late grain development.