[Fluorescence Quantitative PCR Detection of Kinase Region Mutations].

Objective: To establish a highly sensitive and quantitative detection method for kinase region mutations, provide strong support for the early diagnosis and treatment of chronic myeloid leukemia(CML).

Methods: Sampele from 35 CML patients who were initially tested negative for kinase region mutations by Sanger sequencing were collected. The kinase region mutation was detected by the fluorescence quantitative detection kit of Shanghai Yuanqi Biopharmaceutical Technology Co., Ltd. The mutation rate was analyzed by Ct value method. The relative mutation rate of the final kinase region was determined by dividing the mutation rate by the expression level of the fusion gene.

Results: Among the 35 CML patients initially tested negative for mutations by the Sanger sequencing method, 7 cases of T315I mutation, 2 cases of T315A mutation, 2 cases of Y253H mutation, and 1 cases of E255K mutation after detection of the new method. The relative mutation rates range from 0.1% to 19.42%, which could not be detected by Sanger sequencing method. Subsequently, this method was used to detect the mutation in 126 CML patients, and the positive rate exceeded that of the Sanger sequencing method. The gene expression significantly reduced or negative after adjusting treatment strategy based on the mutation situation.

Conclusion: Compared with Sanger sequencing, fluorescence quantitative PCR has higher sensitivity and can screen for low-frequency kinase mutations in the early stage. Moreover, it can also perform relative quantitative analysis, so the method has good clinical application prospects for detecting mutation.