Cognitive abilities are complex human traits influenced by genetic factors. Brain-derived neurotrophic factor (BDNF), a unique polypeptide growth factor, has an influence on the differentiation and survival of neurons in the nervous system. A single-nucleotide polymorphism (rs6265) in the human gene, resulting in a valine to methionine substitution in the pro-BDNF protein, was thought to associate with psychiatric disorders and might play roles in the individual difference of cognitive abilities. However, the specific roles of the gene in cognition remain unclear. To investigate the relationships between the substitution and cognitive abilities, a healthy population-based study and the PCR-SSCP method were performed. The results showed the substitution was associated with digital working memory (p = 0.02) and spatial localization (p = 0.03), but not with inhibition, shifting, updating, visuo-spatial working memory, long-term memory, and others (p > 0.05) among the compared genotype groups analyzed by general linear model. On the other hand, the participants with BDNFGG had higher average performance in digital working memory and spatial localization than the ones with BDNFAA. The findings of the present work implied that the variation in BDNF might play positive roles in human digital working memory and spatial localization.