Chemicals used in DNA Extraction

List of contents Tris-EDTA (TE BUFFER) Phenol Chloroform Iso-amy Alcohol Iso-propanol Ethanol RNAse CTAB NaCl SDS PVP (polyvinylpyrrolidone) Beta-mercaptoethanol

Chemicals used in DNA ExtractionTris-EDTA

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Tris maintains the pH of the solution. Basically it interacts with the lipo-polysaccharides present on the outer membraneof the cell, which helps to permeabilize the membrane. This effect enhanced with the addition of EDTA (Ethylene Diamine Tetra Acetic acid), a molecule chelating cations like Mg2+. The purpose of TE Buffer is to protect DNA or RNA from degradation.

Phenol

Phenol is used to denature the proteins or it helps to remove non-polar proteins and lipids from the solution.

Chloroform

Chloroform prevents cutting of DNA during isolation. It solubilizes lipids and a lot of protein to remove them from the DNA.

Iso-amyl Alcohol

Iso-amyl Alcohol or isopentanol is used to prevent phosgene (CoCl2) from reaction of chloroform (CH3Cl) with air.

Iso-propanol

Its main role is to precipitate the DNA, by engaging with the H2O molecules not giving chance for the DNA to get dissolve in the H2O.

Ethanol

Ethanol is dehydrating agent. In DNA isolation, it is used for the precipitation of DNA molecule. When a molecule is to be precipitated it should be dehydrated as the H2O molecules forming a film around it prevent their interaction.

RNAse

An RNAse is an Enzyme that break down an RNA molecule. RNAses all added to the final solution to ensure that no RNA is in the solution as it may interfere with the DNA in the solution when trying to extract.

CTAB (Cetyl Trimethyl Ammonium Bromide)

CTAB is a best detergent to use during the Extraction/Isolation of highly Polymerized DNA from Plant material. This detergent simultaneously solubilizes the Plant Cell wall and Lipid membranes of internal organelles and denatures proteins (Enzyme). Thus, the DNA is not hydrolysed during the Isolation process and as long as vortexing or vigorous

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shaking are avoided highly Polymerized (i.e. very high intact) genomic DNA is obtained.

NaCl (Salt)

The function of Salt is to precipitate DNA from the solution. The Salt neutralizes the negative charge of the negatively Charged Phosphate in DNA.

SDS (Sodium Dodecyl Sulfate)

Is an Organic Compound, used as detergent that removes the oily strains in the DNA extraction process. It is strong anionic detergent, it removes –ve ions from the protein and destroys its confirmation because of loss of conformation the protein loses its structure.

PVP (polyvinylpyrrolidone)

It helps in removing phenolics by forming hydrogen bonds with them.

Beta-mercaptoethanol

It is added most of the times in the extraction buffers, it is a strong reducing agent to clean tannins and other polyphenols present in the crude plant extract.

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