Question

Explain in brief the separation and isolation of DNA fragments.

Answer 1

Hint: DNA or deoxyribonucleic acid, is a polynucleotide chain polymer which comprises genetic material. The analysis of genetic material gives information on the functioning, reproduction, and growth of an organism.

Complete step-by-step answer:
The analysis of DNA finds use in various fields like paternity testing, criminal investigation, medical diagnosis of diseases, and research.
DNA is obtained from a sample by disrupting the cells. The DNA is separated from the mixture by changing its solubility. It is subjected to the action of the enzyme to form fragments of DNA. The fragmented DNA is subjected to an electric field in an electrophoretic unit and further isolated from the unit for further analysis.
The separation of DNA is done using agarose gel electrophoresis. A low-concentration gel is prepared. The DNA subjected to enzymatic cleavage is loaded into the wells. The gel is subjected to an electric field.
Due to the negative charge on DNA molecules, they move towards the positive electrode. The separation of molecules is dependent on their size. Larger molecules are closer to the well compared to small molecules, which move faster.
The bands are viewed after staining with ethidium bromide under UV light. The band of interest is selected from the gel and cut using a razor.
DEAE membrane is used for transferring the fragments from agarose gel. Using electricity, the fragment is transferred. Further, using salt buffer solutions, the membrane is rinsed followed by extraction using phenol and chloroform.

Note: Various factors like pH, temperature, presence of ions, chain length, incubation time, the volume of EDTA solution, etc affect the stability of DNA.