In this paper, we investigated the dependence of threshold power density on the Al0.55Ga0.45N underlying layer film thickness in ultraviolet-B band (UV-B) lasers on various AlN wafers (four types). We also prepared and compared AlN templates for AlN freestanding substrates, AlN films fabricated by metalorganic vapor phase epitaxy, and annealed sputtered AlN templates at high temperature. The initial growth of AlGaN became three-dimensional by inserting a homoepitaxial Ga-doped AlN layer between the AlN template and Al0.55Ga0.45N, before it shifted to two-dimensional growth. It is possible to reduce the dislocation in Al0.55Ga0.45N using this mode. The dependence of AlGaN film thickness and that of the AlN template on samples with an inserted homoepitaxial Ga-doped AlN layer were studied. Compared with Al0.55Ga0.45N having a thickness of 5 μm, there was almost no noticeable difference between the dark spot density characterized by cathodoluminescence and the threshold power density in UV-B lasers for the AlN template. Besides, the characteristics were noticeably different for the film thickness of Al0.55Ga0.45N. The threshold power density in UV-B laser and dark spot density were reduced by increasing the film thickness. Through the optimization of the crystal growth condition, the threshold power density in UV-B laser and dark spot density were reduced to 36 kW cm−2 and 7.5 × 108 cm−2, respectively.