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Concept
Marqueur de poids moléculaire
Résumé
A molecular-weight size marker, also referred to as a protein ladder, DNA ladder, or RNA ladder, is a set of standards that are used to identify the approximate size of a molecule run on a gel during electrophoresis, using the principle that molecular weight is inversely proportional to migration rate through a gel matrix. Therefore, when used in gel electrophoresis, markers effectively provide a logarithmic scale by which to estimate the size of the other fragments (providing the fragment sizes of the marker are known).
Protein, DNA, and RNA markers with pre-determined fragment sizes and concentrations are commercially available. These can be run in either agarose or polyacrylamide gels. The markers are loaded in lanes adjacent to sample lanes before the commencement of the run.
Although the concept of molecular-weight markers has been retained, techniques of development have varied throughout the years. New inventions of molecular-weight markers are distributed in kits specific to the marker's type.
An early problem in the development of markers was achieving high resolution throughout the entire length of the marker. Depending on the running conditions of gel electrophoresis, fragments may have been compressed, disrupting clarity. To address this issue, a kit for Southern Blot analysis was developed in 1990, providing the first marker to combine target DNA and probe DNA. This technique took advantage of logarithmic spacing, and could be used to identify target bands ranging over a length of 20,000 nucleotides.
There are two common methods in which to construct a DNA molecular-weight size marker. One such method employs the technique of partial ligation. DNA ligation is the process by which linear DNA pieces are connected to each other via covalent bonds; more specifically, these bonds are phosphodiester bonds. Here, a 100bp duplex DNA piece is partially ligated. The consequence of this is that dimers of 200bp, trimers of 300bp, tetramers of 400bp, pentamers of 500bp, etc. will form.
Source officielle
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