Homemade SPRI Bead Recipe

As described in “Towards Large-Scale Museomics Projects: A Cost-Effective and High-Throughput Extraction Method for Obtaining Historical DNA From Museum Insect Specimens” (Holmquist et al. 2025). Modified from Rohland and Reich 2012.

Reagents Required:

  • Cytiva Sera-Mag SpeedBeads™ Carboxyl Magnetic Beads, hydrophobic (Cytiva Supplier #65152105050250)
  • Tris-EDTA (TE) Buffer, or 1M Tris & 500mM EDTA
  • 5M NaCl
  • 1M Tris-HCl
  • 500mM EDTA
  • PEG-8000
  • Tween-20
Reagent Volume (50mL)
Bead Stock Solution 1mL
Tris EDTA (TE) Buffer 50mL
5M NaCl 10mL
1M Tris-HCl 500µL
500mM EDTA 100µL
PEG-8000 10.5 grams
Tween-20 27.5µL

Materials Required:

  • 1.5mL tubes
  • magnet for 1.5mL tubes
  • vortex
  • minicentrifuge
  • pipette
  • scale

Bead Recipe Protocol

  1. Let bead stock come to room temp
  2. Make TE buffer if necessary: Add 500µL 1M Tris and 100µL 500mM EDTA to 50mL falcon tube. Fill to 50mL with diH20.
  3. Homogenize beads
  4. Transfer 1mL of bead stock solution to 1.5mL tube
  5. Place on magnet and let the beads separate, about 5 minutes
  6. Discard supernatant
  7. Add 1mL of TE buffer
  8. Vortex and spin
  9. Place on magnet
  10. Remove TE buffer and let the beads separate, about 5 minutes
  11. Repeat washing with TE buffer (x1)
  12. Repeat washing with TE buffer (x2)
  13. Re-suspend beads in 1mL TE
  14. In 50mL conical, combine 10.5g PEG-8000, 10mL NaCl, 500µL Tris-HCL, 100µL EDTA
  15. Fill up to the ~49mL mark with millipore H20
  16. Vortex until PEG-8000 is fully in solution (takes ~15+ mins)
  17. Add 27.5µL Tween20 and bead solution to conical
  18. Store bead solution at 4ºC, wrap in foil

Testing Your Beads: Ladder Test

Ladder Test Reagents & Materials:

  • GeneRuler Ladder 100bp
  • 6X TriTrack Loading Dye
  • Two sets of 5 microtube strips with lids
  • Millipore water
  • 80% Ethanol (made fresh)
  • Magnet for 0.2mL strip tubes
  • Bead solution

Ladder Test Protocol:

  1. Make fresh 80% ethanol; place in freezer
  2. In a strip of 5 microtubes, aliquot 20µL diluted ladder into each (2µL GeneRuler ladder + 18µL ddH2O, quick vortex and spin)
  3. Homogenize SPRI bead solution, aliquot a small amount (at least 150 µL) into 1.5mL tube and allow to come to room temp
  4. Store bead solution in fridge, wrapped in foil
  5. Add the following volumes of beads based on differing ratios: .8x (16µL), 1.2x (24µL), 1.5x (30µL), 1.8x (36µL), and 2x (42µL)
  6. Incubate 5 mins
  7. Place on magnet and remove supernatant
  8. Add ~80µL 80% ethanol (enough to fully cover beads)
  9. Incubate on magnet (1 minute)
  10. Remove ethanol
  11. Repeat ethanol wash
  12. Ensure no ethanol remains; use single pipette to remove excess
  13. Rehydrate with 20µL of H2O
  14. Briefly vortex the tubes to homogenize, then quick spin
  15. Place on magnet
  16. Transfer supernatant to new tubes for each ratio
  17. Run gel