An open-source algorithm for rapid unbiased determination of DNA fiber length |
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| Affiliation: | 1. Centre de Recherche de l’Hôpital Maisonneuve-Rosemont, Montréal, Québec, Canada;2. Programmes de Biologie Moléculaire, Université de Montréal, Montréal, Québec, Canada;3. Département de Génie Informatique et Génie Logiciel, Polytechnique Montréal, Québec, Canada;4. Département de Médecine, Université de Montréal, Montréal, Québec, Canada;5. Département d’Ophtalmologie, Université de Montréal, Montréal, Québec, Canada;1. Department of Life Science and Biotechnology, Jadavpur University, Kolkata, India;2. PG Department of Microbiology, Bidhannagar College, EB-2 Sector-1, Saltlake, Kolkata, India;3. Rajiv Gandhi Centre for Biotechnology,Thiruvananthapuram 695 014, Kerala, India;1. Centre de recherche de l’Hôpital Maisonneuve-Rosemont, 5415 boulevard de l’Assomption, Montreal, H1T 2M4, Canada;2. Institute for Research in Immunology and Cancer, Université de Montréal, P.O. Box 6128, Succursale Centre-Ville, Montreal, H3C 3J7, Canada;3. Département de Pathologie et Biologie Cellulaire, Université de Montréal, 2900 Edouard Montpetit Blvd, Montreal, H3T 1J4, Canada;4. Département de Médecine, Université de Montréal, Université de Montréal, 2900 Edouard Montpetit Blvd, Montreal, H3T 1J4, Canada;1. Department of Molecular, Cell and Cancer Biology, University of Massachusetts Medical School, Worcester, MA 01605, USA;2. Department of Biology/VGN Proteomics Facility, University of Vermont, Burlington, VT 05405, USA;3. Institute of Genetics, Biological Research Center, Hungarian Academy of Sciences, Szeged 6726, Temesvari krt. 62, Hungary;1. Institute of Molecular Cancer Research, University of Zurich, 8057 Zurich, Switzerland;2. Department of Genetics and Development, Columbia University Irving Medical Center, Irving Cancer Research Center, New York, NY 10032, USA;3. Department of Biochemistry, Vanderbilt University School of Medicine, Nashville, TN 37205-0146, USA;4. Division of Tumor Biology and Immunology, the Netherlands Cancer Institute, 1066 CX, Amsterdam, the Netherlands;1. Department of Genetics, University of North Carolina, Chapel Hill, Genetic Medicine Building 5061, CB#7264, 120 Mason Farm Road, Chapel Hill, NC 27599-7264, USA;2. Curriculum for Bioinformatics and Computational Biology, University of North Carolina, Chapel Hill, 120 Mason Farm Road, Chapel Hill, NC 27599-7264, USA;3. Department of Biochemistry and Biophysics, University of North Carolina, Chapel Hill, 120 Mason Farm Road, Chapel Hill, NC 27599-7264, USA;4. Lineberger Comprehensive Cancer Center, University of North Carolina, Chapel Hill, 120 Mason Farm Road, Chapel Hill, NC 27599-7264, USA;1. Department of Chemical and Systems Biology, Stanford University School of Medicine, Stanford, CA 94305, USA |
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| Abstract: | DNA fiber fluorography is widely employed to study the kinetics of DNA replication, but the usefulness of this approach has been limited by the lack of freely-available automated analysis tools. Quantification of DNA fibers usually relies on manual examination of immunofluorescence microscopy images, which is laborious and prone to inter- and intra-operator variability. To address this, we developed an unbiased, fully automated algorithm that quantifies length and color of DNA fibers from fluorescence microscopy images. Our fiber quantification method, termed FiberQ, is an open-source image processing tool based on edge detection and a novel segment splicing approach. Here, we describe the algorithm in detail, validate our results experimentally, and benchmark the analysis against manual assessments. Our implementation is offered free of charge to the scientific community under the General Public License. |
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| Keywords: | DNA fiber fluorography Automatic Fiber Quantification Open-source fiber analysis |
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