Objective To explore the effects of CD44 gene knockout on behavioral abilities in mice by animal behavior experiment. Methods Beam balance experiment, rotarod experiment, open field experiment, elevated plus-maze experiment, tail suspension experiment, forced swimming experiment, novel object recognition experiment, three-chamber social experiment and Morris water maze experiment were carried out to analyze and compare the behavioral differences between homozygous C57BL/ 6J CD44 knockout (CD44 KO) mice and wild type (WT) mice. Results In Stage 3 of the three-chamber social experiment, WT female mice were seen to spend more time exploring novel objects than female mice in the CD44 KO group (P= 0. 0475). From day 5 to day 7 in the water maze navigation experiment, mice in CD44 KO group were observed to take longer to reach the target area than WT mice (P = 0. 0147, 0. 0182, 0. 0233). In the water maze space exploration experiment, mice in the CD44 KO group tended to reach the target area fewer times (P= 0. 0128) and took longer time to reach the target area for the first time (P= 0. 0003). The time that males in the CD44 KO group took to reach the target area for the first time was less than that taken by females (P= 0. 049). Additionally, males in the CD44 KO group took more time than WT males (P= 0. 0137), and females in the CD44 KO group also took more time than WT females (P= 0. 0036). There were no significant differences in the result of the beam balance experiment, rotarod experiment, open field experiment, elevated plus maze experiment, tail suspension experiment, forced swimming experiment and novel object recognition experiment between the two groups. Conclusions Compared with WT mice,CD44 KO mice were observed to have significantly decreased learning and memory ability, especially in the 5th~ 7th day.In addition, their spatial memory ability was obviously weakened. Meanwhile, CD44 KO female mice were seen to have weaker social propensity than WT female mice. However, no significant differences were noted between CD44 KO mice and WT mice in the aspects of sports coordination ability, balance ability, endurance, anxiety and depression.