Physical mapping involves determining the locations of identifiable landmarks on DNA molecules, such as restriction enzyme cutting sites or genes, and measuring their distances in base pairs. One physical mapping method is fluorescent in situ hybridization (FISH), which detects the positions of markers or genes on chromosomes by hybridizing fluorescent probes to chromosomal DNA on slides under a microscope. Another major approach is restriction fragment overlapping based physical mapping using bacterial artificial chromosomes (BACs). BAC-based physical mapping does not require chromosome slide preparation like FISH and can map hundreds or thousands of genes to contigs.

1. Construction of
Physical mapping

2. Construction of Physical
mapping
• Physical map is the map of the locations of identifiable landmarks on
DNA molecules (e.g. restriction enzyme cutting sites, genes),
regardless of inheritance.
• Distance is measured in base pairs.
• In situ hybridization (ISH) or fluorescent in situ hybridization (FISH) is
one of the physical mapping methods.
• This technique is used to detect the positions of the markers or
genes on chromosomes.
• The fluorescent labeled probes are hybridized to chromosomal DNA
on the slides.
• The position of the fluorescence labeled probes is visualized on the
chromosomes using the microscope or other methods (Jin and Lloyd
1997).
• The major approach to physical mapping is restriction fragment
overlapping based method.

3. In situ hybridasation
• In situ hybridization (ISH) is a type of hybridization that uses a
labeled complementary DNA, RNA or modified nucleic acids
strand (i.e., probe) to localize a specific DNA or RNA sequence
in a portion or section of tissue (in situ).
In molecular biology, a hybridization probe is a fragment
of DNA or RNA of variable length (usually 100–1000 bases long)
which can be radioactively labeled.
It can then be used in DNA or RNA samples to detect the
presence of nucleotide sequences (the DNA target) that are
complementary to the sequence in the probe.

4. DNA probe

5. FISH
• Fluorescence in situ hybridization (FISH) is
a cytogenetic technique that uses fluorescent probes that bind
to only those parts of the chromosome with a high degree of
sequence complementarity.
• It was developed by biomedical researchers in the early
1980s and is used to detect and localize the presence or
absence of specific DNA sequences on chromosomes.
• Fluorescence microscopy can be used to find out where the
fluorescent probe is bound to the chromosomes.

6. FISH

7. Construction of Physical
mapping
• Currently, the BAC-based physical mapping is probably the
most popular method currently used.
• Compared with FISH, BAC-based physical mapping does not
require the preparation of the metaphase chromosome slides.
• Also BAC-based physical mapping can map hundreds or
thousands of genes to the contigs using BES or hybridization
on BAC library.
• These genes will be located on the chromosomes once the
physical map is integrated with the linkage map and the
karyotypes.
• FISH will never provide the same level of resolution as BAC-
based physical mapping.

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