A procedural assessment method suitable for comprehensive and research-oriented Microbiology Experiment course and mixed teaching method has been established. The system involved: (1) online course assessment, including 10 experiment unit tests and the experimental theory test at the end of term. The test score of online course assessment accounts for 20% of the total assessment results. (2) Implement the high-level assessment to cultivate the student’s comprehensive ability and thinking. The first assessment is to make the four experimental reports of module I into the scientific research posters; the second assessment is to write the five experimental reports of module II into scientific research paper; The third assessment is to integrate the three experimental reports of module III into a water quality inspection report; and the fourth is to carry out artistic creation of microbial petri dishes competition. Higher-ranking performance accounted for 30% of the total score. (3) In the final experimental operation test, students randomly selected two exam topics and operated on-site, and all teachers scored simultaneously. The average score of the operational test was accounted for 50% of the total score. The practical operation examination can promote the undergraduate students to master the basic experimental skills of Microbiology. The process assessment of Microbiology Experiment course attaches great importance to the mastery of students’ basic experimental skills and the training of students’ comprehensive ability and thinking mode, which helps to improve the training quality of undergraduate talents in biology science.