Abstract: To establish suitable two-dimensional electrophoresis proteome analysis system (2-D) of melon (Cucumis melo L) leaves,In this experiment,we used melon leaves as experimental material, comparing two different extraction methods leaf protein, and optimized two-dimensional electrophoresis of two important aspects of technology that is focused on the parameters and sample volume.The results showed that: compared with Tirs-phenol extraction, using an improved method Mg/NP-40/PEG3350/TCA acetone extracting leaves protein,and after SDS-PAGEF analyzed, The protein content is higher, Rubisco enzyme compared removed good, bands were clear. When using pH3 ~ 10,18 cm IPG strips of two-dimensional electrophoresis, increasing the low voltage focusing time, get a more clear 2-D map. The protein sample volume 800ug, the maximum number of points were obtained in protein, low protein kurtosis clearer. This experiment established suitable for the melon leaf protein analysis system of two-dimensional electrophoresis, got more than number of protein and high resolution two-dimensional electrophoresis.For the next step in the protein group level analysis of melon disease resistance, yield, properties and related protein provides technical support.