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Lettuce canopy spectral reflectance was measured by hand-held portable spectrum analyzer, the reflectance characteristics was analyzed in the different nitrogen levels and different growth. After the first derivative treatment, then correlation analysis, lettuce canopy nitrogen associated with the four-sensitive band, which were 400 ~ 470nm, 510 ~ 590nm, 600 ~ 670nm, 680 ~ 750nm. Further used stepwise regression analysis, selected the sensitive wavelengths in different growth stages. Partial least squares regression was used for setting up prediction models in different growth. 24 samples were verified models in the same period, the measured and predicted of the correlation coefficient were 0.8570, 0.8853 and 0.8591, and RMSE were 0.6355, 0.6122 and 0.5472.