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    DNA Extraction Reagents - Functions

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    harpreet157
    Plant cell walls and membranes are broken down through grinding and homogenization in an extraction buffer containing Tris, EDTA, sodium chloride, mercaptoethanol, and SDS in order to damage the cell wall, inhibit DNases, disrupt membranes, and release nucleoplasm and DNA. DNA is then purified from other cellular components and impurities through precipitation using potassium acetate and cold ethanol washes followed by centrifugation to obtain a hydrated DNA pellet.

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    DNA Extraction Reagents - Functions

    Uploaded by

    harpreet157
    11K views1 page
    Plant cell walls and membranes are broken down through grinding and homogenization in an extraction buffer containing Tris, EDTA, sodium chloride, mercaptoethanol, and SDS in order to damage the cell wall, inhibit DNases, disrupt membranes, and release nucleoplasm and DNA. DNA is then purified from other cellular components and impurities through precipitation using potassium acetate and cold ethanol washes followed by centrifugation to obtain a hydrated DNA pellet.

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    function of reagents used in plant DNA extraction

    Original Title

    DNA Extraction Reagents- Functions

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    Plant cell walls and membranes are broken down through grinding and homogenization in an extraction buffer containing Tris, EDTA, sodium chloride, mercaptoethanol, and SDS in order to damage the cell wall, inhibit DNases, disrupt membranes, and release nucleoplasm and DNA. DNA is then purified from other cellular components and impurities through precipitation using potassium acetate and cold ethanol washes followed by centrifugation to obtain a hydrated DNA pellet.

    Copyright:

    Attribution Non-Commercial (BY-NC)
    Download as DOC, PDF, TXT or read online from Scribd
    Download as doc, pdf, or txt
    11K views1 page

    DNA Extraction Reagents - Functions

    Uploaded by

    harpreet157
    Plant cell walls and membranes are broken down through grinding and homogenization in an extraction buffer containing Tris, EDTA, sodium chloride, mercaptoethanol, and SDS in order to damage the cell wall, inhibit DNases, disrupt membranes, and release nucleoplasm and DNA. DNA is then purified from other cellular components and impurities through precipitation using potassium acetate and cold ethanol washes followed by centrifugation to obtain a hydrated DNA pellet.

    Copyright:

    Attribution Non-Commercial (BY-NC)
    Download as DOC, PDF, TXT or read online from Scribd
    Download as doc, pdf, or txt
    of 1

    Plant cells posses rigid cellulose cell wall and a cell membrane.

    Breaking the cell, layer by layer


    releases the cellular constituents. At first, the plant cell wall has to be damaged. It is
    accomplished by grinding the plant tissues and homogenizing in the extraction buffer. The
    components of the extraction buffer cause cell lysis.

    Tris (Tris-hydroxymethyl aminomethane) in the extraction buffer, buffers the pH of the


    cells at 8.0. EDTA (Ethylene diamine tetraacetic acid) deters the DNase activity, chelates the
    metal ions which are cofactors for them. It also weakens the cell membrane stability. Sodium
    chloride also helps in maintaining the osmoticum of the cells. Chloride component enter into the
    cells by ionic transport along with water by diffusion. This influx expands the cells, prevents cell
    aggregation and weakens the membrane integrity.

    Mercaptoethanol cleaves the disulphide bridges of proteins and helps in denaturation of


    membrane proteins and cytosolic proteins. SDS is an anionic detergent, which also denatures the
    membrane proteins and disrupts the cell membrane. This also results in the break down of the
    nuclear envelope and thereby releases the nucleoplasm. SDS also helps in inhibition of
    nucleases.
    DNA in the nucleoplasm is neutralized by Potassium acetate. K+ ions bind with negative
    phosphate backbone of the DNA and shield them. This favors DNA precipitation from ethanol in
    cold condition. Upon centrifugation, the neutralized DNA with impurities goes into the
    solution this time. When alcohol is added in cold condition to the solution, DNA strands come
    close together and coalesce. But other impurities are soluble in ethanol and are removed in the
    supernatant. DNA precipitate is settled down as a pellet by centrifugation, purified by 70 %
    ethanol wash, hydrated in TE buffer and analyzed on agarose gel.

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