Abstract: In order to achieve a higher protein discovery rate, many measures were taken to capture as much peptides as possible during the analysis of a practical biological sample. To study the contribution of gradient elution time, repeated runs and mass ranges in the analytical process to the identification rates of proteome in a sample by a shotgun method, yeast lysate was used as a model sample， and a series of experiments were performed on a nano-scale capillary reversed-phase chromatography interfaced to LCQ mass spectrometer to compare the effect between them on achieving the completeness of proteome. The results show that totally identified unique peptides, which finally result in the identification of protein groups, increased remarkably with longer gradient time, repeated runs of a sample and segmented mass ranges, but employing segmented mass ranges is more effective than using longer gradient time or simply repeated runs for improving coverage of yeast protein identifications. The conclusion also suggests that employing complementary analytical strategies should be a better choice in a large-scale proteomic analysis.