DGGE

thumb | Průběh DGGE a CG svorka A very sensitive technique for searching for mutations is electrophoresis in a gradient denaturing gel (denaturing gradient gel electrophoresis, DGGE). It uses the fact that the rate of DNA denaturation DNA depends on the number of hydrogen bonds. DNA strands will separate more easily at sites rich in AT pairs, while stretches rich in CGs will be more stable. A polyacrylamide gel with a gradually increasing concentration of denaturing substances (formamidu a močoviny) elektroforézu (formamidu a močoviny). The examined DNA travels in the electric field at a speed corresponding to its molecular weight, until the moment when the two strands begin to separate from each other. The resulting denatured strands travel much more slowly during electrophoresis, so the easier a certain section is denatured, the closer to the start the DNA sample stops. Since completely separated ssDNA would create fuzzy bands, primers with a so-called CG-clamp are used for DNA amplification. PCR produkt pak obsahuje dvoušroubovice, které mají na jednom konci samé CG páry; v tomto místě se řetězce denaturují jen nesnadno. Citlivost DGGE se blíží 100 %.

TGGE
A similar technique is TGGE (temperature gradient gel electrophoresis, gelová elektroforéza s teplotním gradientem). Instead of a gel with a gradually increasing concentration of denaturing substances, gradually increasing gel temperatures are used.