Studies conducted over the past three decades have revealed strong biological bases for homosexuality and transsexuality.The objectives of this research were to validate the Biological Knowledge of Homosexuality andTranssexuality (BKHT) questionnaire in a sample of students from the area of physical activity in Mexico and describe this knowledge in the sample evaluated.To this end, 324 students from Mexico were surveyed using a sociodemographic survey and the BKHT questionnaire.The results reveal adequate indices of validity and reliability of the BKHT questionnaire in the sample, that the levels of biological knowledge on these topics are low and that the sociodemographic variables that affect these results are sex and having a friend/a known/a homosexual or bisexual. It is recommended to continue with the evaluation of samples of students from other pedagogy careers and from different regions of Mexico.