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Southern Hybridization

Southern blotting is a technique used to detect specific DNA sequences. It involves separating DNA fragments by size through agarose gel electrophoresis, then transferring the DNA to a membrane for probing with labeled DNA. The technique was developed in the 1970s by Edwin Southern and can locate a particular gene within a genome. Key steps are digesting DNA, separating fragments by size, transferring to a membrane, and probing the membrane to identify sequences complementary to the probe.

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5K views17 pages

Southern Hybridization

Southern blotting is a technique used to detect specific DNA sequences. It involves separating DNA fragments by size through agarose gel electrophoresis, then transferring the DNA to a membrane for probing with labeled DNA. The technique was developed in the 1970s by Edwin Southern and can locate a particular gene within a genome. Key steps are digesting DNA, separating fragments by size, transferring to a membrane, and probing the membrane to identify sequences complementary to the probe.

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samdhi
Copyright
© Attribution Non-Commercial (BY-NC)
We take content rights seriously. If you suspect this is your content, claim it here.
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A

PRESENTATION
ON
SOUTHERN BLOTTING

TECHNIQUE

BY: Aakanksha jain


SOUTHERN HYBRIDIZATION
 It is a method routinely used in
molecular biology for detection of a
specific DNA sequence in DNA
samples.
 Southern blotting combines
agarose gel electrophoresis for size
separation of DNA with methods to
transfer the size separated DNA to a
filter membrane for probe
hybridization.
 Southern blotting method is named
after its inventor, the British biologist
Edwin Southern who developed this
procedure at Edinburgh University in
the 1970s.
 Southern blotting could be used to
locate a particular gene within an
entire genome
Steps involved in southern
blotting-

 1. Digest the DNA with an appropriate


restriction enzyme.
 2. Run the digest on an agarose gel.
 3. Denature the DNA (usually while it
is still on the gel).
 4. Transfer the
denatured DNA to the
membrane.
Traditionally, a
nitrocellulose membrane
is used.
 5.Probe the membrane
with labeled ssDNA.
 6.Visualize radioactively
labeled target sequence.
Result-

 Hybridization of the probe


to a specific DNA fragment
on the filter membrane
indicates that this fragment
contains DNA sequence
that is complementary to
the probe.
POLYMERASE CHAIN
REACTION-

 . PCR is the amplification of a


small amount of DNA into a
larger amount. It is quick,
easy, and automated. Larger
amounts of DNA mean more
accurate and reliable results
for your later techniques
 The techniques was developed
by Nobel Laureate Biochemist
Kary Mullis in 1984 and is based
on the discovery of the biological
activity at high temperatures of
DNA Polymerases found in
Thermophiles (bacteria that live in
hot springs).
To perform PCR on a sample
we will need four things-

 1.The target sample


 2.A primer
 3.Taq polymerase
 4. Nucleotides
There are three major steps to PCR
and they are repeated over n over
again -
 1. Target sample is heated
 2. Temperature is reduced
and the primer is added.
 3. New pieces of ssDNA are
made.
Applications of PCR-

 Isolation of genomic
DNA
 Amplification and
quantitation of DNA
 PCR in diagnosis of
diseases
 The Southern blotting technique is
extremely sensitive. It can be used to
map the restriction sites around a
single copy gene sequence in any
genome (even of man). It is used for
DNA fingerprinting, preparation of
RFLP maps, detection and
identification of the transferred genes
in transgenic individuals, etc.
 THANKS

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