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• Always use a flask 2–4 times the volume of the solution. Allow the powder to hydrate in the solution for a few minutes before heating. Weigh the flask and solution before and after heating and add warm distilled water to solution to bring it to its original weight.
• To avoid bubble formation, cool the hot agarose solution to 50–60oC and pour carefully into gel cassette. Allow gel solution to cool gradually; rapid cooling will cause an irregular gel matrix and band distortion during electrophoresis.
• When using low melting agarose gel, it is important to run the gel in a cold buffer. High voltage can cause overheating of the buffer, which can melt the gel. The UltraPure™ Agarose products are made from the highest purity biochemicals and packaged in a pouch that makes pouring and dispensing the powder easier—reducing the likelihood of spills and contamination.

Typically, the TAE buffer provides better resolution of fragments greater than 4 kb, and the TBE buffer resolves 0.1–3 kb fragments. The TBE buffer is better suited for highvoltage (>150V) electrophoresis because of its higher buffering capacity and lower conductivity. If you mistakenly use water instead of buffer, there will essentially be no migration of DNA in the gel. Avoid mishaps with conveniently packaged, ready-to-use 10x TBE or 10x TAE Buffer.

For example, if nucleases are present in PCR reaction, they will effectively cleave any DNA, including the primers and probes. Amplification will therefore not occur. How pure is the source of water used in your PCR reaction? Why not eliminate possible contaminates in your PCR reaction by trying UltraPureTM DNase/RNase Free Distilled Water? With no DNase, RNase, or protease activity detected, it’s designed for use in all molecular biology applications.

The recommended loading volume is dependent on detection methods: Typically, one ng DNA band can be detected with ethidium bromide staining. However, we normally load different volumes—e.g., 10 ul and 2 ul of a DNA ladder in the first and last lanes of a gel to orient the gel and to increase the quantity range. Use of two tracking dyes in the DNA ladders and samples ensures the DNA bands will not run off the gel and indicate maximum resolution has been achieved. Life Technologies offers a wide range of DNA ladders in the ready-to-load TrackItTM format. The TrackItTM DNA Ladders are ready to use—no need to heat, mix, or dilute.

It fluoresces under UV light when the dye intercalates with DNA or RNA. Short exposure of nucleic acids to UV light can cause significant damage to the sample, which will reduce the efficiency of subsequent manipulations of samples, such as ligation or cloning.

If DNA is to be used after separation on agarose gel, it is best to avoid exposure to UV light by using a blue light excitation source and stain, such as SYBR Safe DNA Stain. SYBR Safe DNA Stain is a highly sensitive stain for visualization of DNA that has been shown to have a low level of mutagenicity and toxicity. It is classified as non-hazardous waste with similar sensitivity as EtBr.