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HEINZ-ULRICH G. WEIER
well as detection reagents, and thus increases the hybridization efficiency.
Furthermore, if the DNA molecules can be stretched in some way, they
may provide linear templates for visual mapping. FISH applied in the
past to various types of crude DNA preparations allowed visualization
of probe overlap and it provided some information about the existence
and size of gaps between clones. However, none of those techniques provided sufficiently accurate information about the extent of clone overlap
or the separation between elements in the map because the chromatin
onto which clones were mapped was condensed to varying degrees
from site to site.
We demonstrated previously that cloned DNA fragments can readily
be mapped by FISH onto DNA molecules prepared by the hydrodynamic
action of a receding meniscus and, referring to its quantitative nature, we
termed the technique 'Quantitative DNA Fiber Mapping (QDFM)' (Weier
et al. 1995). In QDFM, a solution of DNA molecules is placed on a glass or
mica surface prepared so that some DNA molecules attach at one or both
ends. The DNA solution is then spread over a larger area by placing a
coverslip on top, and additional DNA molecules are allowed to bind to
the surface. During drying, the molecules are straightened and uniformly
stretched by the hydrodynamic action ofthe receding meniscus. Molecules
prepared in this manner are stretched with remarkable homogeneity. A
properly stretched molecule should extend about -2.3 kb/Jlm, Le., approximately 30% over the length predicted for a double stranded DNA
molecule of the same size (Weier et al. 1995). QDFM can be applied to
DNA molecules ranging in size from a few kb to more than 1 Mbp, which
allows mapping of small probes with near kilobase resolution onto entire
yeast chromosomes and large (mega)YAC clones (Wang et al. 1996, Duell
et al. 1997).
Applications of QDFM extend beyond map assembly and can provide
valuable information for quality control, clone validation, definition of a
minimal tiling path as well as for the sequence assembly process. Furthermore, due to its high hybridization efficiency obtained with DNA fibers,
QDFM is also the method of choice for high resolution optical mapping of
expressed sequences in genomic intervals defmed by the DNA fibers.
HEINZ-ULRICH G. WEIER
well as detection reagents, and thus increases the hybridization efficiency.
Furthermore, if the DNA molecules can be stretched in some way, they
may provide linear templates for visual mapping. FISH applied in the
past to various types of crude DNA preparations allowed visualization
of probe overlap and it provided some information about the existence
and size of gaps between clones. However, none of those techniques provided sufficiently accurate information about the extent of clone overlap
or the separation between elements in the map because the chromatin
onto which clones were mapped was condensed to varying degrees
from site to site.
We demonstrated previously that cloned DNA fragments can readily
be mapped by FISH onto DNA molecules prepared by the hydrodynamic
action of a receding meniscus and, referring to its quantitative nature, we
termed the technique 'Quantitative DNA Fiber Mapping (QDFM)' (Weier
et al. 1995). In QDFM, a solution of DNA molecules is placed on a glass or
mica surface prepared so that some DNA molecules attach at one or both
ends. The DNA solution is then spread over a larger area by placing a
coverslip on top, and additional DNA molecules are allowed to bind to
the surface. During drying, the molecules are straightened and uniformly
stretched by the hydrodynamic action ofthe receding meniscus. Molecules
prepared in this manner are stretched with remarkable homogeneity. A
properly stretched molecule should extend about -2.3 kb/Jlm, Le., approximately 30% over the length predicted for a double stranded DNA
molecule of the same size (Weier et al. 1995). QDFM can be applied to
DNA molecules ranging in size from a few kb to more than 1 Mbp, which
allows mapping of small probes with near kilobase resolution onto entire
yeast chromosomes and large (mega)YAC clones (Wang et al. 1996, Duell
et al. 1997).
Applications of QDFM extend beyond map assembly and can provide
valuable information for quality control, clone validation, definition of a
minimal tiling path as well as for the sequence assembly process. Furthermore, due to its high hybridization efficiency obtained with DNA fibers,
QDFM is also the method of choice for high resolution optical mapping of
expressed sequences in genomic intervals defmed by the DNA fibers.