STEM education has gained popularity in leading economies over the past two decades, but limited research exists on the readiness of in-service and pre-service biology teachers to implement STEM education. This study thus intends to fill this gap by examining the level of Technological Pedagogical Content Knowledge (TPACK) of biology teachers in the implementation of STEM education, with an emphasis on the differences between pre-service and in-service teachers. Quantitative methods employing a questionnaire based on the framework of STEM-TPACK were utilized to evaluate their readiness systematically. The t-test was conducted to analyze the data collected from the sample of 70. Through the lens of the STEMTPACK framework, the findings showed that in terms of content knowledge, pedagogical knowledge, pedagogical content knowledge, technological pedagogical content knowledge, and overall scores, in-service teachers outperformed pre-service teachers significantly. However, pre-service teachers exhibited greater potential in technological knowledge compared to in-service teachers, suggesting the need for further investigation. Using Guangdong, the most populous province of China, as the target district, this study provided an understanding of the readiness of pre-service and in-service biology teachers in implementing STEM education respectively. The next phase of this research aims to explore the underlying reasons and provide detailed insights into the quantitative results, which will be presented in another paper.