Understanding phylogenetic trees representing evolutionary relationships of living organisms is essential in school biology. Traditional instructions based on lectures and textbooks with pictorial presentations have been proven ineffective to promote students’ understanding of the concept. This study, therefore, developed a game-based learning activity called the VERT card game to enhance students’ learning of the phylogenetic taxonomy of chordates. The VERT card game was designed to lay the foundation for different characteristics of chordate classes, as well as to allow students to construct and interpret their evolutionary relationships based on the phylogenetic tree. The effectiveness of the VERT card game was verified by a pre-test and post-test design with 109 middle school students in Thailand. The statistical result revealed that students’ mean scores increased significantly in the post-test, compared to the pre-test, indicating their improved understanding. In addition, after participating in the learning activity, the student participants were found to increase their self-efficacy to learn evolutionary biology statistically. Also, it showed positive views towards the usefulness of the developed card game as a large number of them expressed that they would like to use it for their lesson reviews and wished to have this form of learning activity in other topics in biology.