The uncertainty is a necessary part of the measurement results. Given the coordinate data and uncertainty of the measured discrete points, the computing procedure and method of uncertainty in straightness error assessment were discussed. In the study, the measured point coordinates were considered to conform to uniform distribution. Based on the decision rules of minimum condition, the uncertainty property of the reference line and its solution were proposed. Then the transmission formulas and computing method of uncertainty were given，and Monte Carlo method was applied to get the straightness error and uncertainty. Finally, an example was given to verify the method, and the non-uniform distribution of the measured points was discussed.