Linear DNA can be resolved by size using agarose gels of various concentrations. The greater the percentage of agarose, the smaller the linear DNA that can be resolved. The sugar polymers that make up the agarose gel matrix (powdered agarose heated in appropriate buffer, poured into a gel tray and allowed to solidify) act like a sieve. The greater the agarose concentration, the smaller the pores created in the gel matrix, and the more difficult it is for large linear DNA molecules to move through the matrix. Changing the agarose concentration changes the size of the sieve matrix of the gel. However, there is an upper and lower limit to accurate separation of DNA molecules using agarose gel electrophoresis. To estimate the size of a linear DNA fragment and to diagnose any electrophoresis issues, run at least one DNA molecular weight marker [e.g., BenchTop 1kb DNA Ladder (Cat.# G7541)] along with the experimental sample. A table highlighting the various agarose percentages and molecule sizes that can be separated is shown below.
Resolution of Linear DNA on Agarose Gels.
| Recommended % Agarose | Optimum Resolution for Linear DNA |
|---|
| 0.5 |
1,000–30,000bp |
| 0.7 |
800–12,000bp |
| 1.0 |
500–10,000bp |
| 1.2 |
400–7,000bp |
| 1.5 |
200–3,000bp |
| 2.0 |
50–2,000bp |
For standard analytical gels, we recommend an agarose like Agarose, LE, Analytical Grade (Cat.# V3121). For preparative gels, where fragments are to be separated and cut from a gel, a low-melting-point agarose such as Agarose, LMP, Preparative Grade for Large Fragments (>1,000bp; Cat.# V2831) or Agarose, LMP, Preparative Grade for Small Fragments (10 to 1,000bp; Cat.# V3841) is advantageous.