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Quantitative DNA Fiber Mapping

  • Lu Chun-Mei
  • Mei Wang
  • Karin M. Greulich-Bode
  • F. Weier Jingly
  • G. Weier Heinz-UlliEmail author
Protocol
  • 1.6k Downloads
Part of the Springer Protocols Handbooks book series (SPH)

Several hybridization-based methods that are used to delineate single-copy or repeated DNA sequences over larger genomic intervals take advantage of the increased resolution and sensitivity of free chromatin, i.e., chromatin released from interphase cell nuclei. Quantitative DNA fiber mapping (QDFM) differs from the majority of these methods in that it applies FISH to purified, clonal DNA molecules which have been bound to a solid substrate at one end (at least). The DNA molecules are then stretched by the action of a receding meniscus at the water–air interface, which results in the DNA molecules being stretched homogeneously to about 2.3 kb/µm. When nonisotopically, multicolor-labeled probes are hybridized to these stretched DNA fibers, and their respective binding sites are visualized under the fluorescence microscope, their relative distances can be measured and converted into kilobasepairs (kb). The QDFM technique has found a variety of useful applications, ranging from the detection and delineation of deletions or overlaps between linked clones, to the construction of high-resolution physical maps and studies of stalled DNA replication and transcription.

Keywords

Yeast Artificial Chromosome Yeast Artificial Chromosome Clone Maleic Acid Buffer Alkaline Lysis Protocol Alkaline Lysis Solution 
These keywords were added by machine and not by the authors. This process is experimental and the keywords may be updated as the learning algorithm improves.

Abbreviations

BAC

Bacterial artificial chromosome

CEPH

Centre des Études du Polymorphisms Humain, Paris, F

LMP

Low melting point

PCR

Polymerase chain reaction

PFGE

Pulsed field gel electrophoresis

QDFM

Quantitative DNA fiber mapping

UTR

Untranslated region

YAC

Yeast artificial chromosome

Notes

Acknowledgments

This work was supported by a grant from the Director, Office of Energy Research, Office of Health and Environmental Research, US Department of Energy, under contract DE-AC02-05CH11231.

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Copyright information

© Springer-Verlag Berlin Heidelberg 2009

Authors and Affiliations

  • Lu Chun-Mei
    • 1
  • Mei Wang
    • 2
  • Karin M. Greulich-Bode
    • 3
  • F. Weier Jingly
    • 4
    • 5
  • G. Weier Heinz-Ulli
    • 6
    Email author
  1. 1.E.O. Lawrence Berkeley National Laboratory (LBNL), Life Sciences DivisionUniversity of CaliforniaBerkeleyUSA
  2. 2.US DOE Joint Genome Institute/E.O. LBNLWalnut CreekUSA
  3. 3.Deutsches Krebsforschungszentrum (DKFZ), DKFZ/ZMBH AllianzAbteilung Genetik der HautkarzinogeneseHeidelbergGermany
  4. 4.Life Sciences DivisionUniversity of California, E.O. LBNLBerkeleyUSA
  5. 5.Reprogenetics, LLCSouth San FranciscoUSA
  6. 6.Life Sciences Division, MS 74-157University of California, E.O. LBNLBerkeleyUSA

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