With a view of detecting the effects of macromolecular crowding on the phase transition of DNA compaction con-fined in spherical space, Monte Carlo simulations of DNA compaction in free space, in confined spherical space without crowders and in confined spherical space with crowders were performed separately. The simulation results indicate that macromolecular crowding effects on DNA compaction are dominant over the roles of multivalent counterions. In addition, effects of temperature on the phase transition of DNA compaction have been identified in confined spherical space with different radii. In confined spherical space without crowders, the temperature corresponding to phase transition depends on the radius of the confined spherical space linearly. In contrast, with the addition of crowders to the confined spherical space, effects of temperature on the phase transition of DNA compaction become insignificant, whereas the phase transition at different temperatures strongly depends on the size of crowder, and the critical volume fraction of crowders pertains to the diameter of crowder linearly.