As a typical dual-phase titanium alloy, Ti-6Al-4V has a complex phase transformation process, which is sensitive to temperature parameters and difficult to control its microstructure. To further master the law of phase transformation of Ti-6Al-4V titanium alloy and make a reasonable heat treatment system, different heat treatment systems in β single-phase region and (α+β) two-phase region were made for Ti-6Al-4V titanium alloy, and the phase transformation process and microstructure growth rate of different heat treatment processes were studied by high-temperature confocal microscope. At 1050 °C and 1200 °C in the β single-phase region, the β grain size increases with the prolongation of the holding time and the increase of the holding temperature. The tissue growth rate of the α phase was higher than 1200°C in the cooling process after 1050°C incubation, which was 2.88 μm/s. Based on the heat preservation in the single-phase region, the second heat preservation was carried out for the two-phase region of Ti-6Al-4V titanium alloy at 850 °C and 900 °C. The study found that when the holding temperature was 850°C, the fastest growth rate of the α phase was 4.64 μm/s, which was greater than 3.29 μm/s when the holding temperature was 900°C.