Agarose gel electrophoresis is a fundamental technique in molecular biology, widely used to separate DNA fragments based on their size. It involves casting a gel from agarose, a polysaccharide derived from seaweed, which forms a porous matrix. When an electric current is applied, negatively charged DNA molecules migrate through these pores, with smaller fragments moving faster and further than larger ones. This separation allows researchers to visualize, analyze, and even purify specific DNA fragments.
Key Storage Practices
Proper storage maintains the integrity and usability of agarose gels. The optimal temperature for storing cast agarose gels is 4°C, in a refrigerator. This cool temperature slows degradation and inhibits microbial growth.
To prevent dehydration and contamination, gels should be stored in an airtight manner. This involves wrapping them tightly in plastic wrap or placing them in sealed containers or plastic bags. If stored for more than a day, gels can be kept moist by adding a small amount of the same buffer used to cast them. For gels with light-sensitive stains, storing them in the dark preserves the stain’s effectiveness. Labeling the stored gel with its pour date and concentration tracks its age and composition.
Factors Influencing Gel Longevity
The type of buffer used to prepare the agarose gel affects its stability. Tris-acetate-EDTA (TAE) and Tris-borate-EDTA (TBE) are common electrophoresis buffers. TAE buffer has lower buffering capacity and is more susceptible to microbial contamination, shortening gel shelf life. TBE buffer, with its boric acid, offers higher buffering capacity and greater stability, but borate can interact with DNA and can precipitate at low temperatures during long-term storage.
DNA-binding stains also impact gel longevity and storage. Stains like ethidium bromide (EtBr) or SYBR Safe are light-sensitive and degrade when exposed to light, diminishing DNA visualization. Microbial contamination poses a threat to gel integrity, particularly in gels prepared with TAE buffer; proper sealing and refrigeration reduce this risk. Over extended periods, agarose itself can undergo chemical degradation through hydrolysis, breaking down the gel matrix.
Recognizing Gel Degradation
Several visual cues indicate a degraded agarose gel. Evaporation is a common issue, leading to shrunken, brittle, or cracked gels. This results from inadequate sealing during storage. Visible mold or bacterial colonies signal microbial contamination.
Changes in the gel’s clarity or texture suggest degradation of the agarose matrix. Using degraded gels negatively impacts electrophoresis results, leading to poor band resolution, smeared DNA bands, or distorted migration patterns. For instance, degraded DNA appears as a smear rather than distinct bands. Gels exhibiting these signs should be discarded for reliable experimental outcomes.