Selecting the structure with the first two clustering results, used Discovery Studio to remove backbone atoms, and non-target bases, and then added the hydrogen atom on the N1 position of the pyrimidine base and the N9 position of the purine base. The remaining residues were as the structure for running quantum chemistry calculations. Single-point quantum chemistry calculations were run on the above structure in Orca 4.1.2 ( wB97X-D3 level of theory, def2-TZVP basis set, and def2-TZVP/C auxiliary basis set. The solvation model was the conductor-like polarized continuum model, and the solvent probe radius was 1.5 Å to overcome the inconsistency caused by implicit solvent calculations. For the energy of each stack, there were six interactions: two base pairing hydrogen bond energies (𝐸 HB ), two intra-stand stack energies and two inter-strand stack energies (𝐸 stack ). For the energy of each dangle, there were two interactions: an intra-stand stack energy and an inter-strand stack energy. Interactions energies were calculated by the balance method of Boys and Bernardi which minimizes the basis set superposition error.