Table of Contents
Why was salt used in the extraction solution?
Salt is used during the process of DNA extraction because salt helps in neutralizing DNA molecules, which makes the DNA molecule less hydrophilic, i.e. less soluble in water. Salt also helps in detaching protein molecules bound to the DNA. By which, the extraction process will be easier.
What is the role of NaCl in CTAB buffer?
NaCl: Helps to remove proteins that bind to the DNA and keep the proteins dissolved in the aqueous layer, so they do not precipitate in the alcohol along with the DNA by neutralizing the negative charges (Heikrujam et al., 2020).
How does salt concentration affect DNA?
Experimental as well as theoretical results show that the DNA molecule is more stable as the concentration of salt (or cations) increases. It is known that the two strands of DNA molecule carry negative charge due to phosphate group along the strands.
Why is protease used in DNA extraction?
Proteases catalyze the breakdown of contaminating proteins present in the solution to its component amino acids. It also degrades any nucleases and/or enzymes that may be present in the sample. This is of vital importance since these chemical compounds can attack and destroy the nucleic acids in your sample.
What is the role of isoamyl alcohol in DNA extraction?
Isoamyl alcohol: In the phenol-chloroform DNA extraction method, Isoamyl alcohol helps in reducing foaming between interphase. It prevents the emulsification of a solution. The liquid phase contains DNA and the organic phase contains lipid, proteins and other impurities.
What is the molarity of NaCl used during the process of DNA extraction?
So for NaCl concentrations under 0.5M, CTAB and DNA molecules can create complexes. In those concentrations, proteins and other hydrocarbons are still soluble in water except the DNA-CTAB complex.
How does nacl prevent denaturation of DNA?
It is known that the two strands of double stranded DNA(dsDNA) have negative charge due to phosphate group. Cations in the form of salt in the solution, act as shielding agents thereby reducing the repulsion between these strands.
Can nacl denature DNA?
DNA Denaturation through Salt A high concentration of salt will cause DNA to naturally denature, given the right concentration of salt. DNA denaturation with salts are similar to denaturation through the use of organic solvents. In general, DNA denaturation through salt cannot be renatured.
Why is isopropanol used in DNA extraction?
The overall function of salt and ethanol/ isopropanol is to precipitate DNA from the solution. The salts neutralize the negative charge of the negatively charged phosphate in DNA and the isopropanol /ethanol removes the hydration shell of H2O molecules around the phosphate.
What is the role of phenol in DNA extraction?
Extraction of DNA containing samples with acidic phenol results in the denaturation of the DNA, and once denatured, the DNA partitions to the organic phase. This is a key feature of many RNA purification protocols, which is one of the reasons acidic buffer-saturated phenol is used.
What is the role of beta mercaptoethanol in DNA extraction?
Plants are rich in phenolics compounds and to get a quality DNA these should be removed. β-Mercaptoethanol (HOCH2CH2SH) is added most of the time in extraction buffers and is a strong reducing agent to clean tannins and other polyphenols present in the crude plant extract. Globular proteins get dissolved in water.