Write any four methods to get free pure D.N.A. Explain
Answers
Answer:
Some of the most common DNA extraction methods include organic extraction, Chelex extraction, and solid phase extraction.[3] These methods consistently yield isolated DNA, but they differ in both the quality and the quantity of DNA yielded. When selecting a DNA extraction method, there are multiple factors to consider, including cost, time, safety, and risk of contamination.
Organic extraction involves the addition of and incubation in multiple different chemical solutions;[3] including a lysis step, a phenol chloroform extraction, an ethanol precipitation, and washing steps. Organic extraction is often used in laboratories because it is cheap, and it yields large quantities of pure DNA. Though it is easy, there are many steps involved, and it takes longer than other methods. It also involves the unfavorable use of the toxic chemicals phenol and chloroform, and there is an increased risk of contamination due to transferring the DNA between multiple tubes.[4] Several protocols based on organic extraction of DNA were effectively developed decades ago,[5] though improved and more practical versions of these protocols have also been developed and published in the last years.[6]
Chelex extraction method involves adding the Chelex resin to the sample, boiling the solution, then vortexing and centrifuging it. The cellular materials bind to the Chelex beads, while the DNA is available in the supernatant.[4] The Chelex method is much faster and simpler than organic extraction, and it only requires one tube, which decreases the risk of DNA contamination. Unfortunately, Chelex extraction does not yield as much quantity and the DNA yielded is single-stranded, which means it can only be used for PCR-based analyses and not for RFLP.[4]
Solid phase extraction such as using a spin-column based extraction method takes advantage of the fact that DNA binds to silica. The sample containing DNA is added to a column containing a silica gel or silica beads and chaotropic salts. The chaotropic salts disrupt the hydrogen bonding between strands and facilitate binding of the DNA to silica by causing the nucleic acids to become hydrophobic. This exposes the phosphate residues so they are available for adsorption.[7] The DNA binds to the silica, while the rest of the solution is washed out using ethanol to remove chaotropic salts and other unnecessary constituents.[3] The DNA can then be rehydrated with aqueous low salt solutions allowing for elution of the DNA from the beads.
Answer:
Four steps are used to remove and purify the DNA from the rest of the cell.
Lysis.
Precipitation.
Wash.
Resuspension.