Biochemical Techniques for the Extraction of Escherichia Coli Genomic DNA

Biochemical Techniques for the Extraction of Escherichia Coli Genomic DNA

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Modern biochemical study and analysis of nucleic acids have been heavily dominated by electrophoresis and polymerase chain reaction techniques, as the former allows for relatively inexpensive and accessible resolution and visualization of nucleic acids according to basic chemical properties such as molecular charge and weight, and the latter quickly increases the concentration of nucleic acids, normally found in cells in minute amounts, to a level easily analyzed by modern biochemical techniques. These two techniques are therefore currently indispensable in dealing with nucleic acids on a practical level, and are tools which should be present in every biologist’s kit. This study therefore attempts to elucidate the theoretical and practical processes involved in both the polymerase chain reaction and agarose gel electrophoresis through the extraction and visualization of Escherichia coli genomic DNA through said techniques.

Nucleic acids are necessary biomolecules as they function in the expression, transmission and storing of genetic information needed for the synthesis of proteins. They are responsible for passing on genetic information from one cell generation to the succeeding cell generation. These nucleic acids can be broken down into nucleotides, which are composed of a 5-carbon ribose sugar, an amine or nitrogenous base and a phosphate. The ribose sugars can be used to classify the two types of nucleic acids; namely DNA (deoxyribonucleic acid) and RNA (ribonucleic acid). The amines are classified as either a purine or pyrimidine. The purines are guanine and adenine while the pyrimidines are cytosine, thymine and uracil.

Deoxyribonucleic acid or DNA is a double helix composed of a pentose sugar and phosphate backbone ...

... middle of paper ...

...NA fragments will consequently have an easier time moving through the gel mesh thus larger fragments will produce bands near the top of the gel while the smaller fragments can be found at the bottom.

Finally, the DNA can be visualized through staining and subsequent UV illumination. Two staining methods can be used, the first is rapid DNA staining through BioRad Fast Blast DNA stain solution, the second is through the use of ethidium bromide which is more sensitive than the first but is more dangerous since the main solution is a carcinogenic material.

In this experiment, however, restriction enzymes were not used such that the results showed little success, as judged by the lack of DNA bands in the agarose gels that were used to check the results sue to the isolated genomic DNA being so large that it does not migrate in the gel and instead remains in the well.

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