High-throughput approach for detection of DNA bending and flexibility based on cyclization.
We have developed a high-throughput approach to the labor-intensive problems of DNA cyclization, which we use to characterize DNA curvature and mechanical properties. The method includes a combinatorial approach to make the DNA constructs needed and automated real-time measurement of the kinetics using fluorescence. We validated the approach and investigated the flexibility of two kinds of nicked DNA and AT dinucleotide repeats. We found that, although the nicks hardly alter the bending flexibility, they significantly increase the torsional flexibility, and that the AT repeat has 28% (+/-12%) lower bending rigidity than a generic DNA sequence.
Zhang
Yongli
Y
Departments of Molecular Biophysics and Biochemistry and Chemistry, Yale University, New Haven, CT 06511, USA.
Crothers
Donald M
DM
D001483:Base Sequence; D004247:DNA / Q000138:chemical synthesis / Q000737:chemistry*; D011088:DNA Ligases; D018900:Dinucleotide Repeats; D015195:Drug Design; D005456:Fluorescent Dyes; D015394:Molecular Structure; D009690:Nucleic Acid Conformation; D012235:Rhodamines; D013050:Spectrometry, Fluorescence
C114557:5-carboxytetramethylrhodamine succinimidyl ester; D005456:Fluorescent Dyes; D012235:Rhodamines; D004247:DNA; D011088:DNA Ligases