[agarose on scale] As mentioned in the text on the previous page, more agarose means a firmer gel, and a more difficult matrix for the DNA to move through. We can use this information to optimize the experimental conditions if we have a general idea how large our DNA fragments will be.

For example, gels are often mixed at 0.8-1.0% agarose. This concentration will separate a wide range of fragment sizes, ranging from around 500 base pairs (bp) to around 20,000 bp (or 20 kilobases, kb). Separation simply means being able to differentiate between 2 or more fragments of different sizes.

If less agarose is used (e.g., 0.5%), fragments less than around 2 kb will all be equally challenged by the agarose matrix and will migrate equal distances in equal time, providing no separation. Large fragments (e.g., >20 kb), however, will be able to separate better in this matrix than in a 1.0% gel, where they would be blocked from moving by all the agarose around.

Conversely, if more agarose is used (e.g., 2.0%), better separation of very small fragments will be seen, because even very small fragments will be challenged by the high level of agarose. However, at this high concentration, anything larger than 3-4 kb will not separate because there is just too much agarose for those fragments to move efficiently.

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