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Importance of gel immersion in the running buffer A gel must be fully submerged in running buffer with 3–5 mm of buffer covering the gel’s surface. Insufficient running buffer can cause poor resolution, band distortion, or even melting of the gel. However, excess running buffer will also decrease DNA mobility and cause band distortion. How agarose gels work When set, an agarose gel will form a complex polysaccharide matrix. You can think of this matrix as a cheese grater (stay with me) which provides resistance against nucleic acid molecules. Taking the cheese grater analogy, if you want large pieces of grated cheese you would use a grater with larger holes in it. On the other hand, if you only want tiny pieces of cheese shavings you would use a grater with very fine holes. This same concept occurs within the agarose gel matrix. The higher the concentration of agarose within the gel, the smaller the pores within the matrix (or holes in a cheese grater). These smaller pores make it difficult for larges molecules to migrate through the gel and so they favour the smaller fragments in the sample. Conversely, lower concentration agarose gels have larger pores which allow the separation of larger fragments easily. So, when a current is applied through the gel during electrophoresis, the nucleic acids migrate through the pores and are separated based on their molecular size. It is the size of these pores which ultimately decides the molecule sizes can pass through the agarose gel. #NikolaysGeneticsLessons #gelElectrophoresis #gelElectrophoresisExplained #gelElectrophoresisOfDna #gelElectrophoresisProcedure #gelElectrophoresisLab #gelElectrophoresisAnalysis #agaroseGelElectrophoresis #agaroseGelElectrophoresisProtocol #agaroseGelElectrophoresisOfDna #agaroseGelElectrophoresisExplained #dnaGelElectrophoresis #dnaElectrophoresis #electrophoresisOfDna #sdsPageGelElectrophoresis #polyacrylamideGelElectrophoresis #glycerol #TBE
