Introduction: The medical profession is one of the most demanding disciplines and historically remained a male-dominated profession. Academic performance in medical science is affected by various sociodemographic factors. Various studies have been done to predict academic performance in medical school regarding the influence of gender and attendance. In this study, an attempt was made to study the effect of gender and attendance on performance in formative assessment in the subject of biochemistry. Aims and objectives: To compare performance in theory, practical, and combined (theory and practical) formative assessments between male and female students in the subject of biochemistry in the first-year MBBS and to compare class attendance in theory, practical, and combined (theory and practical) between male and female students in the subject of biochemistry in the first-year MBBS. Materials and methods: This study was carried out at the Department of Biochemistry, GMERS Medical College, Himmatnagar. Students’ academic records in the subject of biochemistry during their first professional year of MBBS were studied. For the study, data of 450 (251 male and 198 female) students from 3 academic years were collected. Performance in theory and practical formative assessments was compared between male and female students. A comparison of marks of male and female students was carried out using Student's t-test. The effect of attendance on performance was assessed gender-wise. For comparison of attendance, Kruskal–Wallis test was applied. For all comparisons, a p value <0.05 was considered significant. All statistical analyzes were carried out using Microsoft Excel 2010. Results: The performance of female students was better than male students in theory and combined formative assessments (p < 0.05). The attendance of female students in theory classes and combined attendance of theory and practical was significantly higher (<0.05) than their male counterparts. In different attendance groups for both male and female students, the performance was significantly better with more attendance (p < 0.05). Discussion: In our study, we explored how gender and attendance can affect the performance of the student in the subject of biochemistry. Our study demonstrates female students have more class attendance and better performance in formative assessment in the subject of biochemistry and both the genders’ performances are positively correlated with their attendance.