Gel Electrophoresis Calculator
Calculate expected migration distance and DNA/protein band positions on agarose or PAGE gels.
Why Migration Is Log-Linear
A gel is a porous matrix, and molecules move through it by threading between pores. Larger molecules encounter more obstruction, and the relationship between size and mobility turns out to be logarithmic rather than linear.
This is why size standards are plotted as log(size) against distance to give a straight line, and why gels resolve small differences well at one end of the range and poorly at the other. A 100 bp difference is obvious at 500 bp and invisible at 10 kb.
| Agarose % | Resolution range (bp) |
|---|---|
| 0.5% | 1,000–30,000 |
| 0.8% | 800–12,000 |
| 1.0% | 500–10,000 |
| 1.5% | 200–3,000 |
| 2.0% | 50–2,000 |
| 3.0% | 10–500 |
Higher percentage means smaller pores, resolving small fragments better while large ones fail to enter and pile up near the well. Matching gel percentage to the expected fragment range is the single most important choice in setting up a run.
Why SDS-PAGE Works at All
Native proteins have varied charge and shape, so they would migrate unpredictably. SDS solves this by coating every protein with negative charge in proportion to its length — roughly one SDS molecule per two residues — so charge-to-mass ratio becomes constant and separation depends on size alone.
| Acrylamide % | Protein range (kDa) |
|---|---|
| 7.5% | 50–200 |
| 10% | 20–150 |
| 12% | 10–70 |
| 15% | 5–50 |
When Apparent Size Misleads
| Situation | Effect on apparent mass |
|---|---|
| Glycosylation | Runs larger — sugars add mass and drag |
| Highly acidic proteins | Often run larger — bind less SDS |
| Incomplete denaturation | Compact form runs smaller |
| Supercoiled plasmid | Runs faster than linear of same size |
The plasmid case catches people out routinely. Uncut plasmid gives multiple bands — supercoiled, nicked circular and linear forms all migrate differently despite identical mass. Supercoiled runs fastest because it is most compact.
Worked Examples
Common Mistakes
Large fragments fail to enter a high-percentage gel and small ones run off a low-percentage one. Match the gel to the expected size range.
The log-linear relationship holds only within the resolving range. Extrapolating beyond the smallest or largest marker band is unreliable.
Glycosylated proteins run larger, and highly acidic ones often do too because they bind less SDS. Discrepancies of 10–20% are common.
Supercoiled, nicked and linear forms of the same plasmid migrate differently. Three bands from one plasmid is normal.