Measuring DNA Hybridization Kinetics in Live Cells Using a Time-Resolved 3D Single-Molecule Tracking Method
| Autor: | Pengyu Ren, Thomas E. Yankeelov, Angela Liu, Huong T. Vu, Stephanie Phillion, Yu-An Kuo, Hsin-Chih Yeh, Soonwoo Hong, Yin-Jui Chang, Trung D. Nguyen, Yen-Liang Liu, Yuan-I. Chen, Cong Liu |
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| Rok vydání: | 2019 |
| Předmět: |
Time Factors
Kinetics 010402 general chemistry 01 natural sciences Biochemistry Phase Transition Article Catalysis Diffusion chemistry.chemical_compound Colloid and Surface Chemistry Mammalian cell Molecule Measurement method Chemistry DNA–DNA hybridization Temperature Nucleic Acid Hybridization DNA General Chemistry Markov Chains Single Molecule Imaging In vitro 0104 chemical sciences Solutions Biophysics Nucleic acid Nucleic Acid Conformation Algorithms |
| Zdroj: | Journal of the American Chemical Society. 141:15747-15750 |
| ISSN: | 1520-5126 0002-7863 |
| Popis: | Single-molecule detection enables direct characterization of annealing/melting kinetics of nucleic acids without the need for synchronization of molecular states, but the current experiments are not carried out in a native cellular context. Here we describe an integrated 3D single-molecule tracking and lifetime measurement method that can follow individual DNA molecules diffusing inside a mammalian cell and observe multiple annealing and melting events on the same molecules. By comparing the hybridization kinetics of the same DNA strand in vitro, we found the association constants can be 13- to 163-fold higher in the molecular crowding cellular environment. |
| Databáze: | OpenAIRE |
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