Fast and quantitative differentiation of single-base mismatched DNA by initial reaction rate of catalytic hairpin assembly.
The widely used catalytic hairpin assembly (CHA) amplification strategy generally needs several hours to accomplish one measurement based on the prevailingly used maximum intensity detection mode, making it less practical for assays where high throughput or speed is desired. To make the best use of the kinetic specificity of toehold domain for circuit reaction initiation, we developed a mathematical model and proposed an initial reaction rate detection mode to quantitatively differentiate the single-base mismatch. Using the kinetic mode, assay time can be reduced substantially to 10 min for one measurement with the comparable sensitivity and single-base mismatch differentiating ability as were obtained by the maximum intensity detection mode. This initial reaction rate based approach not only provided a fast and quantitative differentiation of single-base mismatch, but also helped in-depth understanding of the CHA system, which will be beneficial to the design of highly sensitive and specific toehold-mediated hybridization reactions.
Beijing National Laboratory for Molecular Sciences (BNLMS), Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Institute of Analytical Chemistry, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China.
Chenxi
C
Li
Beijing National Laboratory for Molecular Sciences (BNLMS), Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Institute of Analytical Chemistry, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China.
Yixin
Y
Li
Beijing National Laboratory for Molecular Sciences (BNLMS), Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Institute of Analytical Chemistry, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China.
Xiao
X
Xu
Department of Chemical Biology, School of Pharmaceutical Sciences, Peking University, Beijing 100083, China.
Xinyi
X
Wang
Beijing National Laboratory for Molecular Sciences (BNLMS), Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Institute of Analytical Chemistry, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China.
Yang
Y
Chen
Department of Chemical Biology, School of Pharmaceutical Sciences, Peking University, Beijing 100083, China.
Xiaoda
X
Yang
Beijing National Laboratory for Molecular Sciences (BNLMS), Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Institute of Analytical Chemistry, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China.
Feng
F
Liu
Beijing National Laboratory for Molecular Sciences (BNLMS), Key Laboratory of Bioorganic Chemistry and Molecular Engineering of Ministry of Education, Institute of Analytical Chemistry, College of Chemistry and Molecular Engineering, Peking University, Beijing 100871, China. Electronic address: lina@pku.edu.cn.
Na
N
Li
D020137:Base Pair Mismatch / Q000235:genetics*; D001483:Base Sequence; D002384:Catalysis; D004247:DNA / Q000737:chemistry / Q000235:genetics*; D004252:DNA Mutational Analysis / Q000379:methods*; D055029:Inverted Repeat Sequences / Q000235:genetics*; D007700:Kinetics; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques / Q000379:methods*; D015203:Reproducibility of Results; D012680:Sensitivity and Specificity; D016415:Sequence Alignment / Q000379:methods*; D017422:Sequence Analysis, DNA / Q000379:methods*
D004247:DNA
Catalytic hairpin assembly; DNA; Initial reaction rate; Single-base mismatch