Direct and precise length measurement of single, stretched DNA fragments by dynamic molecular combing and STED nanoscopy
A combination of DNA stretching method and super-resolution nanoscopy allows an accurate and precise measurement of the length of DNA fragments ranging widely in size from 117 to 23,130 bp. BstEII- and HindIII-treated λDNA fragments were stained with an intercalating dye and then linearly stretched on a coverslip by dynamic molecular combing. The image of individual DNA fragments was obtained by stimulated emission depletion nanoscopy. For DNA fragments longer than ∼1000 bp, the measured lengths of DNA fragments were consistently within ∼0.5 to 1.0 % of the reference values, raising the possibility of this method in a wide range of applications including facile detection for copy number variations and trinucleotide repeat disorder.
KeywordsDNA Dynamic molecular combing Length measurement STED nanoscopy
We thank Professor Stefan W. Hell of Max Planck Institute for Biophysical Chemistry for the permission to use the Imspector imaging program. This research was supported by the Global Frontier Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Science, ICT & Future Planning (NRF-2014M3A6A4075063). We also acknowledge the BK21 Plus Program and SNU Brain Fusion Grant.
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Conflict of interest
The authors declare that they have no conflict of interest.
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