DNA: 20% PEG 8000 + 2.5 M NaCl + 10 mM Tris–HCl + 1 mM EDTA + 0.05% Tween 20, pH 8.0 @ 25 °C
RNA: 20% PEG 8000 + 2.5 M NaCl + 1 mM citrate + 0.05% Tween 20, pH 6.4 @ 25 °C
Purpose
- SPRI bead mix, such as AMPure XP and RNAClean XP, is an alternative to spin columns for purifying nucleic acids out of aqueous solutions.
- After the DNA/RNA is purified on the beads, they can be resuspended in a reaction mix and left in during the reaction ("with-bead protocol"); afterward, this buffer will rebind them to the same beads, saving on the cost of bead mix (it is equivalent to the buffer used in the bead mix).
- This recipe includes Tween 20 to reduce surface tension and bead clumping.
- The DNA version uses TE to protect DNA from degradation; the RNA version uses citrate.
Recipe (DNA version)
- Final concentrations
| PEG 8000
|
20% (w/v)
|
| Sodium chloride
|
2.5 M
|
| Tris base
|
10 mM
|
| Disodium EDTA
|
1 mM
|
| Tween 20
|
0.05% (v/v)
|
| HCl
|
3.36 mM
|
- Preparing 50 mL of 1X working solution
| Sodium chloride, 5 M
|
25.000 mL
|
| Nuclease-free water
|
3.582 mL
|
| Tris base, 1 M
|
0.500 mL
|
| Disodium EDTA, 0.1 M
|
0.500 mL
|
| HCl, 1 N
|
0.168 mL
|
| PEG 8000, 50% (w/v)
|
20.000 mL
|
| Tween 20, 10% (v/v)
|
0.250 mL
|
Recipe (RNA version)
- Final concentrations
| PEG 8000
|
20% (w/v)
|
| Sodium chloride
|
2.5 M
|
| Trisodium citrate
|
1 mM
|
| Tween 20
|
0.05% (v/v)
|
| HCl
|
0.56 mM
|
- Preparing 50 mL of 1X working solution
| Sodium chloride, 5 M
|
25.000 mL
|
| Nuclease-free water
|
4.672 mL
|
| Trisodium citrate, 1 M
|
0.050 mL
|
| HCl, 1 N
|
0.028 mL
|
| PEG 8000, 50% (w/v)
|
20.000 mL
|
| Tween 20, 10% (v/v)
|
0.250 mL
|
Notes
- HCl titration was calculated with the Python package ionize 0.8.0.
- pH is temperature-dependent and this dependency is especially strong for Tris, so pH indicators may show a noticeably higher pH than 8.0 at temperatures lower than 25 °C.