An operation method of a non-volatile memory for reducing the second-bit effect in the non-volatile memory is suitable for an N-level memory cell having a first storage position and a second storage position (wherein N is a positive integer greater than 2). The method includes following steps: determining sets of operation levels for operating the first storage position according to the level of the second storage position; when the level of the second storage position is a lower level, operating the first storage position according to a first set of operation levels; when the level of the second storage position is a higher level, operating the first storage position according to a second set of operation levels. Each of the levels in the second set of operation levels is greater than the corresponding level in the first set of operation levels.