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Fluorescence in situ Hybridization (FISH)

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Fluorescence in situ
Hybridization
Fluorescence in situ
Hybridization (FISH)
пЃ· FISH
- a process which vividly
paints chromosomes or portions
of chromosomes with
fluorescent molecules
пЃ· Opening picture - Human Mphase spread using DAPI stain
Fluorescence in situ
Hybridization (FISH)
пЃ· Identifies
chromosomal
abnormalities
пЃ· Aids in gene mapping,
toxicological studies, analysis of
chromosome structural
aberrations, and ploidy
determination
Fluorescence in situ
Hybridization (FISH)
пЃ· Used
to identify the presence
and location of a region of DNA
or RNA within morphologically
preserved chromosome
preparations, fixed cells or
tissue sections
Fluorescence in situ
Hybridization (FISH)
пЃ· This
means you can view a
segment or entire chromosome
with your own eyes
пЃ· Was often used during M phase
but is now used on I phase
chromosomes as well
Fluorescence in situ
Hybridization (FISH)
пЃ· Advantage:
less labor-intensive
method for confirming the
presence of a DNA segment
within an entire genome than
other conventional methods like
Southern blotting
FISH Procedure
пЃ· Denature
the chromosomes
пЃ· Denature the probe
пЃ· Hybridization
пЃ· Fluorescence staining
пЃ· Examine slides or store in the
dark
FISH Procedure
FISH Uses
пЃ· Detection
of high
concentrations of base pairs
пЃ· Eg: Mouse metaphase
preparation stained with DAPI
(a non-specific DNA binding dye
with high affinity for A-T bonds)
FISH Uses
FISH Uses
пЃ· Centromere
regions stained
brighter - means they are rich in
A-T bonds
пЃ· Also used in germ cell or
prenatal diagnosis of conditions
such as aneuploidies
FISH and Telomeres
пЃ· Telomeric
probes define the
terminal boundaries of
chromosomes (5’ and 3’ ends)
пЃ· Used in research of
chromosomal rearrangements
and deletions related to cell
aging or other genetic
abnormalities
FISH and Telomeres
пЃ· Special
telomeric probes
specific to individual
chromosomes have been
designed
пЃ· Probe is based on the TTAGGG
repeat present on all human
telomeres
FISH and Telomeres
FISH and Telomeres
пЃ· Application
in cytogenetics can detect submicroscopic
deletions and cryptic
translocations of genes
associated with unexplained
mental retardation and
miscarriages
FISH and Telomeres - Medical
from TelVysion DNA website
 “A translocation
between
chromosomes 12 and 21 with
breakpoints at bands 12p13
(TEL)[telomere] and 21q22
(AML1) occurs in at least 25
percent of childhood B-cell
acute lymphocytic leukemia”
FISH and Telomeres - Medical
пЃ· The
“LS1 TEL/AML1 ES DualColor Translocation Probe” is
designed to detect the TEL and
AML1 gene fusion
пЃ· This fusion is undetectable by
standard cytogenetics, but can
be seen with FISH
FISH - Medical
пЃ· FISH
can be used in the study
of transgenic animals (eg: Polly)
пЃ· Selective markers show if the
human DNA was inserted
successfully and pinpoint where
the human DNA is
пЃ· Transgenic Mouse
FISH - Medical
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