A cost-effective detection of low-abundance mutation with DNA three-way junction structure and lambda exonuclease |
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Authors: | Zishan Feng Wei Zhang Longjie Li Bocheng Tu Weicong Ye Xiaofeng Tang Hongbo Wang Xianjin Xiao Tongbo Wu |
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Affiliation: | 1. School of Pharmacy, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China;2. Institute of Reproductive Health, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China;3. Department of Obstetrics and Gynaecology, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, China |
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Abstract: | We presented a low-abundance mutation detection method with lambda exonuclease and DNA threeway junction structure.The assistant strand in the DNA three-way junction structure could regulate the reaction system from the kinetics and thermodyna mics aspects.The optimization of the assista nt strand helps to improve the selectivity of the mutant-type DNA to the wild-type DNA about 35 times.Moreover,the cost of the optimization process could be saved by about 90%.The method was applied to the detection of a human ovarian cancer-related gene mutation BRCA1(rs1799949,c.2082 CT).The limit of detection to the mutation abundance in the DNA three-way junction structure system(0.2%) was one order lower compared with that in the double-stranded DNA structure system(2%).The mutation abundance in different standard samples was quantitively measured,and the results were consistent with the initial abundance in the standard samples. |
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Keywords: | Gene mutation Cancer Low abundance DNA three-way junction structure Lambda exonuclease |
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