This paper describes a fully autonomous real-time in-motion alignment algorithm for Strapdown Inertial Navigation Systems (SINS) in land vehicle applications. Once the initial position is available, the vehicle can start a mission immediately with accurate attitude, position and velocity information determined within ten minutes. This is achieved by two tightly coupled stages, that is, real-time Double-vector Attitude Determination Coarse Alignment (DADCA) and Backtracking Fine Alignment (BFA). In the DADCA process, the vehicle motion is omitted to roughly estimate the attitude at the very start of the alignment. Meanwhile, attitude quaternions and velocity increments are extracted and recorded. The BFA process utilises the stored data and exploits the Non-Holonomic Constraints (NHC) of a vehicle to obtain virtual velocity measurements. A linear SINS/NHC Kalman filter with mounting angles as extended states is constructed to improve the fine alignment accuracy. The method is verified by three vehicle tests, which shows that the accuracy of alignment azimuth is 0·0358° (Root Mean Square, RMS) and the positioning accuracy is about 15 m (RMS) at the end of the alignment.