Abstract: AIM: To establish a pyrosequencingbased method for simultaneous and rapid detection of CYP2C9^*3 gene polymorphism.METHODS: With the results of standard ABI sequencing as a standard control, we observed and analyzed the reliability of the pyrosequencing method which involves the establishment of biotin-labeled primers, amplification of PCR, separation of Beads, preparation of pyrosequencing single-stranded template of mutational sites and sequencing of PyroMark ID in pyrosequencing apparatus by detected 220 cases of human DNA samples.RESULTS: We achieved the simultaneous and rapid detection platform of CYP2C9^*3 mutation of DNA samples based on Pyrosequencing technology, which can detect DNA samples with high-flux manner.The test results of the CYP2C9^*3 mutation of 96 DNA samples can be obtained each time with this detection platform.Compared with the standard ABI sequencing, the mutation detection rate and the repetition rate of CYP2C9^*3 mutation are 100% in this method.CONCLUSION: Pyrosequencing method is a accurate, high-throughput, rapid method for the CYP2C9^*3 mutation detection, especially suitable for the bulk detection of this mutation.

Key words: pyrosequencing, polymorphism, sequence analysis, CYP2C9