Accomplished physical education (PE) teachers recognize teachers' motivation, lesson implementation as well as the application of PE contents. This is because these affect teaching effectiveness in PE class. However, a previous study from Koonsri (2016) found that the teachers lacked motivation and focused on extra money or bonus from their teaching. In addition, because the pre-service PE teachers had poor experiences in teaching and controlling the classes, these issues affected the implementation and integration of the PE contents (Claypan, 2014; Kanchana, 2010). Thus, this study examines the relationship between teachers' motivation, lesson implementation, and application of PE contents among pre-service PE teachers in Southern Thailand. The instruments regarding teachers' motivation in terms of intrinsic and external motivation, lesson implementation, and application of PE contents were adopted from Tulyakul (2019). The respondents were 157 pre-service PE teachers in five universities in the border provinces of Southern Thailand. The result shows that there is a relationship between intrinsic and external motivation towards lesson implementation and the application of PE contents among the participants. Finally, there are proposed guidelines for the pre-service PE teachers to adjust and develop their teaching performance to enhance students' achievement in the future.