Recommend the optimal DNA ladder, compute loading-dye volumes, estimate gel run time, and generate a well layout for 8/16/24/48-well mini and midi gels.
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Volume of PCR product per well.
Gel loading plan for 12 samples on 1 × 16-well mini gel (1.5% agarose). Per-sample loading mix: • 5.0 µL PCR product • 1.0 µL 6× loading dye • Total per well: 6.0 µL (well capacity: 20 µL) Ladder: 100 bp DNA Ladder (100–1,500 bp), 5 µL per lane, 1 lane per gel. Run conditions: • 5 V/cm × 7 cm = 35 V • Estimated run time: 88 min (until dye front reaches ~70% of gel length) • Check at 44 min for dye front position
When to use
Do not use for
On 24- or 48-well gels, band migration varies by up to 10% from center to edge. Having a ladder at both ends lets you interpolate band sizes more accurately. The calculator auto-adds the second ladder lane for batches over 16 samples.
Standard genotyping gels run at 5 V/cm. You can push to 7-8 V/cm for a faster run, but above 10 V/cm the Joule heating melts low-percentage agarose gels and distorts band resolution. If you need speed, use a high-throughput pre-cast gel system, not higher voltage.
Run time estimates are just that — estimates. The actual stopping point is when the leading dye front (bromophenol blue) reaches 70% of the gel length. Stop there. Running to the bottom wastes time and runs small fragments off the gel.
Ladder recommendation uses a band-size threshold: 100 bp ladder for <1 kb, 1 kb ladder for 1-10 kb, lambda HindIII for >10 kb. Loading dye volume is sample_volume / (6× − 1) = sample_volume / 5 to achieve 1× final concentration from a 6× stock. Run time is estimated from gel length 0.7 (run 70% of gel), divided by a migration rate of 5 cm/hr at 1% agarose and 5 V/cm, scaled inversely by agarose percentage and directly by voltage. Well layout assigns ladder(s) to the first (and optionally last) well, fills sample wells sequentially, and marks remaining wells as empty.
Last validated 2026-04-07. Calculations are designed for planning and documentation support; verify procurement decisions against manufacturer specifications or institutional SOPs.
ConductScience Gel Loading & Ladder Helper (v1.8.0). ConductScience, Inc. 2026. Available at: https://conductscience.com/tools/gel-loading-helper
Lee PY, Costumbrado J, Hsu CY, Kim YH. Agarose gel electrophoresis for the separation of DNA fragments. J Vis Exp. 2012;(62):e3923.
Sambrook J, Russell DW. Molecular Cloning: A Laboratory Manual. 3rd ed. Cold Spring Harbor: CSHL Press; 2001.
DNA is negatively charged at neutral pH. When you apply an electric field, DNA fragments migrate through the agarose matrix toward the positive electrode (anode). Shorter fragments move faster because they encounter less frictional resistance. The agarose concentration sets the pore size — higher concentrations create smaller pores that slow everything down but improve resolution of small fragments.
1. It adds density (glycerol or Ficoll) so the sample sinks into the well instead of floating away. 2. It includes tracking dyes (bromophenol blue, xylene cyanol, or Orange G) that migrate at known rates so you can see when to stop the gel. Bromophenol blue runs at roughly the same speed as a 300 bp fragment in 1% agarose; xylene cyanol runs with ~4 kb fragments.
A DNA ladder is a set of known-size fragments that you run alongside your samples. By comparing your band's position to the ladder, you estimate its size. Choose a ladder whose densest band spacing covers your expected band size — a 100 bp ladder has bands every 100 bp from 100-1500, while a 1 kb ladder has bands at 250, 500, 750, 1000, 1500, 2000, 3000, 4000, 5000, 6000, 8000, and 10000 bp.
Usually means degraded DNA (old proteinase K in the lysis step, or too many freeze-thaw cycles) or overloaded wells. Reduce sample volume or re-lyse with fresh proteinase K. Can also be caused by loading too much loading dye — the glycerol smears if it is >2× the standard 1× concentration.
Caused by uneven heating — the center of the gel gets hotter and the DNA migrates faster there. Lower the voltage or move the gel to a cold room. For wide midi-gels, use a recirculating buffer system.
Check: (1) did you add EtBr or SYBR Safe to the gel or the running buffer? Without a stain you will not see anything. (2) Did the gel run in the right direction? DNA runs toward the red (positive) electrode. (3) Is the PCR actually producing product? Run a positive control.
ConductColony prints gel-loading maps that match your PCR plate map, so every well is pre-labeled with the animal ID and expected genotype.
Open app EquipmentScale the PCR master mix that produces the products you are about to load on this gel.
Browse EquipmentScale the lysis buffer volumes for the tissue that feeds the PCR that feeds this gel.
Browse EquipmentAfter the gel: test whether your observed genotype ratios match expectations.
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