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has emerged form the development in the
area of genetic engineering and molecular
biology.
this technique is used to detect the presence
of DNA from pathogens in clinical specimens
and locate specific genes in cell.
is a method for identifying closely related
nucleic acid molecules within two
populations, a complex target population and
a comparatively homogeneous probe
population
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Definition of nucleic acid hybridization
h. Target DNA
2. Probe
3. Detection System
4. 'ormat
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h. Dot Blot Assay - a specified volume of
sample or specimen is spotted onto a small
area of nitocellulose membrane.
2. Southern Hybridization Assays ² invlove
restriction enzyme digestion and agarose
gel electrophoresis of the target DNA prior
to the hybridization assay.
- the different bands on the agarose gel are
transferred by capillary action onto a nitocellulose
or nylon membrane in a blot apparatus.
Dot blot hybridization analysis of DNA from the LMW and HMW
gel fractions with probes for repetitive sequences known to be present
in T.brucei: subtelomeric repeats (subtel), the conserved second part
of VSG genes (VSG), 70, h77 and V0 bp and rDNA repeats and
kDNA minicircles; hybridization signals are quantitated relative to
non-fractionated control DNA (bloodstream form DNA digest;
tubulin is h).
Southern Hybridization Assays
In Situ Hybridization Assays
- during the transfer, each if the DNA bands is
transferred onto the membrane in the same
relative position that it had in the gel.
3. In Situ Hybridization Assays ² involve
the probing of intact cells or tissue
sections affix to a microscope slides.
- this type of solid phase assay has the advantage
that one cannot only detect the presence of
target DNA in intact cells but also determine
the location of such target DNA within a tissue.
- an important application of in situ
hybridization is for the detection of viruses and
certain types of bacteria within infected cell.
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