Here, we used our model to predict DNA hybridization free energies. To construct training data, a hybridization pool was established, including 10,000 perfect duplexes and 10,000 duplexes with dangles (Figure ). Short duplexes tend to denature at room temperatures, rendering their melt temperatures unmeasurable. Long duplexes may adopt one or more secondary structures in their single strand, and melt temperatures at different concentrations are close or even overlap, leading to biased experimental thermodynamic measurements. Consequently, the length of the stack domain was designed ranging from 5 to 14 nucleotides. Since the free energy of dangles with more than 4 nucleotides can be approximated to that of dangles with 4 nucleotides, the length of the dangle domain was designed ranging from 1 to 4 nucleotides. Given the possibility of dangles positions in duplexes, ten structures were identified in the pool. The longest DNA string contains 49 characters, thus zero-padding was applied to all descriptor matrices. High resolution melt is the optimal method for measuring DNA hybridization thermodynamics since it requires lower DNA concentrations compared to isothermal titration calorimetry and has higher efficiency in data acquisition compared to ultraviolet melt. Thus, high resolution melt analysis was used as the labeling method of the pool, which measured melt curves of duplexes by the real-time fluorescent quantitative PCR (Figure ) and determined hybridization free energies by fitting analysis.