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    Southern Blot, Northern Blotting, Western Blot

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    uziaftab5
    The document discusses Southern blotting and Northern blotting techniques. Southern blotting is used to detect DNA sequences using gel electrophoresis, transfer, and hybridization with a labeled probe. Northern blotting similarly detects RNA sequences. Both techniques allow identification of specific sequences and have applications in research, diagnosis, and forensics.

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    Southern Blot, Northern Blotting, Western Blot

    Uploaded by

    uziaftab5
    19 views18 pages
    The document discusses Southern blotting and Northern blotting techniques. Southern blotting is used to detect DNA sequences using gel electrophoresis, transfer, and hybridization with a labeled probe. Northern blotting similarly detects RNA sequences. Both techniques allow identification of specific sequences and have applications in research, diagnosis, and forensics.

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    A brief explanation of southern blotting and northern blotting.

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    southern blot, northern blotting, western blot ppt

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    The document discusses Southern blotting and Northern blotting techniques. Southern blotting is used to detect DNA sequences using gel electrophoresis, transfer, and hybridization with a labeled probe. Northern blotting similarly detects RNA sequences. Both techniques allow identification of specific sequences and have applications in research, diagnosis, and forensics.

    Copyright:

    © All Rights Reserved
    Download as PPTX, PDF, TXT or read online from Scribd
    Download as pptx, pdf, or txt
    19 views18 pages

    Southern Blot, Northern Blotting, Western Blot

    Uploaded by

    uziaftab5
    The document discusses Southern blotting and Northern blotting techniques. Southern blotting is used to detect DNA sequences using gel electrophoresis, transfer, and hybridization with a labeled probe. Northern blotting similarly detects RNA sequences. Both techniques allow identification of specific sequences and have applications in research, diagnosis, and forensics.

    Copyright:

    © All Rights Reserved
    Download as PPTX, PDF, TXT or read online from Scribd
    Download as pptx, pdf, or txt
    of 18

    Southern blot &

    Northern blot
    Presented To: Dr Raza Hameed
    Presented By: Uzma Batool
    Roll No.05
    MPhil Microbiology
    What is blotting?

     Blots are techniques for transferring


    DNA, RNA and proteins onto a
    carrier so they can be separated,
    and often follows the use of gel
    electrophoresis. The southern blot is
    used to transferring DNA, the
    Northern blot for RNA and the
    western blot for Protein.
    Southern blotting

     This method involves separation, transfer and hybridization.


     The Southern blotting is used to detect the presence of a particular
    piece of DNA in a mixture of sample.
     The DNA detected can be a single gene, or it can be part of larger
    piece of DNA such as viral genome.
     The key to this method is Hybridization.
     HYBRIDIZATION: Process of forming a double stranded DNA molecule
    between a single-stranded DNA probe and a single stranded target
    patient DNA.
    Principle

    1. The mixture of molecule is separated.


    2. The molecules are immobilized on a matrix transferred to a
    solid support (blot).
    3. The labeled probe is added to the matrix to bind to the
    molecule.
    4. Any unbound probes are then removed.
    5. The place where the probe is connected corresponds to the
    location of the immobilized target molecule.
    Steps in Southern blotting

    1. The DNA to be analyzed,


    such as the total DNA of an
    organism, is digested to
    completion with a
    restriction enzyme.
    2. The complex mixture of
    fragments is subjected to
    gel electrophoresis to
    separate the fragment
    according to size.
    Cont.

    3. The restriction fragments present


    in the gel are denatured with heat or
    alkali and transferred onto a
    nitrocellulose or nylon membrane by
    blotting.
    4. This procedure preserves the
    distribution of the fragments in the
    gel, creating a replica of the gel on
    the filter.
    Cont.

    4. The filter is incubated under


    hybridization conditions with a specific
    radio labeled DNA probe.
    5. The probe hybridizes to the
    complementary DNA restriction
    fragment.
    6. Excess probe is washed away and
    the probe bound to the filter is
    detected by autoradiography, which
    reveals the DNA fragment to which the
    probe hybridized.
    APPLICATIONS

     To identify specific DNA in a DNA sample.


     To Isolate desired DNA for construction of rDNA.
     Identify mutations, deletions, and gene rearrangements.
     Used in prognosis of cancer and in prenatal diagnosis of genetic
    diseases.
     Diagnosis of HIV-1 and infectious disease.
     Criminal Identification and Forensics
     Personal Identification
    Northern Blotting

     Northern blotting is a technique for detection of specific


    RNA sequences.
     Northern blotting was developed by Jamse Alwine and
    George Stark at Stanford University.
     The Northern blot procedure essentially identical to that
    of southern blotting except that here RNAs are separated
    by gel electrophoresis.
    Steps involved in Northern blotting

    1. RNA is isolated from


    several biological samples,
    RNA is more susceptible to
    degradation than DNA.
    APPLICATIONS

     A standard for direct study of gene expression at the level


    of mRNA.
     Detection of mRNA transcript size .
     Study RNA degradation.
     Northern blotting has been used in diagnostic settings to
    detect specific RNA markers associated with specific
    disease state, such as viral infections, cancer and genetic
    disorders.
    Thank You

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