Tuesday, May 19, 2020

Extraction of Dna from Calf or Hog Thymus/Isolation of...

Extraction of DNA from Calf or Hog Thymus/Isolation of Yeast RNA I. Abstract Nucleic acids may be divided into two groups RNA and DNA. DNA contains almost all the genetic information while RNA serves as the bridge between the DNA and proteins. Study of both DNA and RNA initially involves proper extraction/isolation. The storehouse of eukaryotic DNA is the nucleus (and in the mitochondria), so experimentally, DNA is extracted from tissues that have a high nuclear to cytoplasmic mass ratio, such as the tissues of the thymus gland and spleen. The thymus gland is a particularly good site for DNA extraction because it functions as the primary site for T lymphocyte differentiation. The T lymphocytes it acts upon have a round nucleus that†¦show more content†¦7.4 (at this pH, stability of crucial hydrogen bonds and other linkages in the DNA double helix are maintained). The addition of sodium citrate buffer has two other significances. First, it chelates Mg2+ and Mn2+, cations that serve as cofactors for DNAses. This chelating action of sodium citrate prevents the degradative enzymes to take effect in performing their function. Secondly, sodium citrate chelates other divalent cations (e.g. Ca2+) that could form salts w ith the anionic phosphate groups in the DNA backbone. Centrifugation was done next in order to separate the nuclei from other cellular organelles as well as cellular debris. The denser nuclei sank down to the bottom after centrifugation, forming the residue, whilst the  ¡Ã‚ §lighter ¡Ã‚ ¨ cellular organelles such as the mitochondria, etc. remained suspended in the supernatant. After discarding off the supernatant, the residue was suspended in 24 ml of 2.0 M NaCl. This was done because the salt increases the ionic strength of the solvent that in turn produces a  ¡Ã‚ §salting in ¡Ã‚ ¨ effect. Salting in then allowed the proteins of the nucleus to again, dissolve for easier separation later through a second centrifugation. Another effect of salt addition is that it could, with strong possibility, weaken the interactions between the negative charges carried by DNA and the positive charge of basic proteins. SDS solution was then

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