Caesium chloride gradient

Plasmid DNA can be purified by the classical alkaline lysis/caesium chloride gradient procedure or through rapid methods involving the use of small amon-exchange resin columns (e.g, Qiagen or Nucleobond columns)... 

The materials usually used to produce such gradients are salts of heavy alkali metals, caesium chloride being the most frequently used. This salt has a high solubility and its low relative molecular mass permits rapid diffusion enabling the gradient to be formed reasonably quickly. Concentrations of salt up to about 2 g ml-1 can be used and are chosen on the basis of information about the density of the test particles or macromolecules. The technique is used frequently in the separation of viral particles and nucleic acids. 

Figure 13.7 Caesium chloride density gradient centrifugation for (a) the separation of DNA from RNA and protein and (b) the separation of linear DNA and supercoiled DNA.

RNA and DNA are isolated from tissues using caesium chloride density gradient sedimentation... 

When total DNA is subjected to high speed density gradient centrifugation in caesium chloride, a large band of nuclear DNA and one or more small satellite bands are usually observed. Up to four satellite DNA bands have been detected in... 

SK 23) (Table 2) using a modified hexadecylt-rimethylammonium bromide (CTAB) extraction protocol (Doyle and Doyle 1990 Lange 1997). Nuclear DNA was purified by ultracentrifugation through a caesium chloride—ethidium bromide density gradient (Lange 1997). This DNA was used to create enriched microsatelhte libraries... 

For zonal density gradient centrifugation there are a number of media that can be used, however the most common of these is sucrose whereas the most common medium for isopycnic density gradient centrifugation is caesium chloride (CSCI2). 

The sedimentation velocity of a molecule becomes zero when the density of the molecule equals the density of its solvent. This condition is readily obtained with DNA molecules in concentrated caesium chloride solutions. If such a solution is spun in an ultracentrifuge at speeds between 25,000 and 50,000 rev/min, the caesium chloride forms a density gradient in the rotor and the DNA molecules move to the region where their density equals that of the solvent. Apparent densities of DNA molecules (around 1.7 g ml in CsCl) can be measured to an accuracy of +0.0002 g mU and preparative... 

The use of caesium sulphate as a gradient-forming salt, instead of caesium chloride, to give improved separations of proteoglycans from bovine nasal cartilage has been advocated. The distribution of the link proteins in different proteoglycan extracts may be followed. 

Gradient-forming materials which provide the densities required for the separation of subcellular particles include salts of alkali metals (e.g. caesium and rubidium chloride), small neutral hydrophilic organic molecules (e.g. sucrose), hydrophilic macromolecules (e.g. proteins and polysaccharides), and a number of miscellaneous compounds more recently introduced and not included in the above group, such as colloidal silica (e.g. Percoll) and non-ionic iodinated aromatic compounds (e.g. Metrizamide, Nycodenz and Renograffin). 

Direction of sedimentation Caesium f chloride J gradient Marker mixture of light and heavy DMAs... 

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