Agarose Gel Calculator
Recommend agarose percentage by DNA size and calculate gel preparation.
Agarose Gel Concentration Reference
| Gel % | Separation range (kb) | Optimal (kb) |
|---|---|---|
| 0.5% | 1–30 kb | 10–30 kb |
| 0.7% | 0.8–12 kb | 5–10 kb |
| 1% | 0.5–10 kb | 3–7 kb |
| 1.2% | 0.4–7 kb | 2–5 kb |
| 1.5% | 0.2–4 kb | 0.5–3 kb |
| 2% | 0.1–3 kb | 0.2–1.5 kb |
| 3% | 0.05–1 kb | 0.1–0.5 kb |
Learn more
What it does
The agarose gel calculator works in two directions: given a target DNA size it recommends a gel percentage and shows the effective separation range, and given a percentage and volume it calculates how much agarose powder and buffer to weigh out. A reference table of concentrations against resolvable size ranges, based on standard molecular cloning practice, is shown alongside. It answers the two questions asked most often at the bench, namely which gel to pour and how much powder to add.
How it works
Agarose forms a hydrated mesh whose pore size shrinks as the concentration rises, and DNA migrating through that mesh moves at a rate that decreases roughly with the logarithm of fragment length. Consequently low percentage gels, around 0.5%, keep large fragments of 1 to 30 kilobases distinguishable, while high percentage gels, up to 3%, resolve small fragments from about 50 to 1000 base pairs; outside its range a gel either lets fragments run together near the front or fails to separate them at all near the well. Gel preparation is a straightforward weight-for-volume calculation: agarose mass in grams = percentage x volume in millilitres / 100, so a 1% gel is 1 gram of agarose per 100 millilitres of running buffer, and the buffer volume is taken as the gel volume since the powder contributes negligible volume.
Worked example
Enter a target size of 0.3 kilobases, that is a 300 base pair colony PCR product, and the calculator recommends a 1.5% gel with an effective separation range of 0.2 to 4 kilobases; if you need to distinguish 300 from 350 base pairs, going to 2% or 3% sharpens resolution further at the small end. Switching to preparation mode with 1.5% and 80 millilitres gives 1.5 x 80 / 100 = 1.2 grams of agarose dissolved in 80 millilitres of 1x TAE or TBE. For the very common 1% 100 millilitre gel, the same arithmetic gives exactly 1 gram.
When to use it
When screening colony PCR products of a few hundred base pairs, a 1.5 to 2% gel keeps the bands well separated from the primer dimer front, whereas the 1% gel used by default would run them close together. When checking a restriction digest of a plasmid with fragments spanning several kilobases, a 0.7 to 1% gel keeps the large fragments resolved instead of stacking them near the well. When you need to cut a band out for extraction, preparation mode tells you the powder and buffer for a thicker, larger-volume gel that tolerates a wide well and a clean excision.
FAQ
- What agarose percentage should I use for a 500 bp fragment?
- A 1.5 to 2% gel is the usual choice, since both concentrations resolve fragments in the low hundreds of base pairs while keeping the run time reasonable. A 1% gel will still show the band clearly, but neighbouring sizes will be harder to distinguish, so go higher whenever you need to tell similar small fragments apart.
- How much agarose powder do I need for a 1% 100 mL gel?
- One gram, because percentage here is weight per volume, so 1% means 1 gram per 100 millilitres. Weigh the powder into the buffer rather than the reverse, heat until fully dissolved with no visible lens-shaped particles, and let the solution cool to about 60 degrees Celsius before adding stain and pouring.
- Why do large DNA fragments run poorly on a high-percentage gel?
- Because the pores are too small for large molecules to thread through efficiently, so fragments above the gel's range all migrate very slowly and pile up as a single poorly resolved band near the well. The fix is a lower percentage gel, and for fragments beyond roughly 30 kilobases a lower voltage with a longer run, or pulsed-field electrophoresis, is needed.
- Should I use TAE or TBE buffer for agarose gels?
- TAE has lower buffering capacity but gives better resolution and mobility for large fragments, and it interferes less with enzymatic steps after gel extraction, which makes it the default for cloning work. TBE buffers more strongly and resolves small fragments better during long or high-voltage runs, but borate can carry over and inhibit downstream enzymes.