This is a cached page for the URL (http://kuchem.kyoto-u.ac.jp/seika/shiraishi/protocols/sds.html). To see the most recent version of this page, please click here.
Protocol Online is not affiliated with the authors of this page nor responsible for its content.
About Cache
SDS-PAGE

SDS-polyacrylamide gel electrophoresis


SOLUTIONS
30% acrylamide mix29% (w/v) acrylamide
1% (w/v) N,N'- methylenebisacrylamide
5x Electrophoresis bufferTris base: 15.1 g
Glycice: 94 g
SDS: 5 g
H2O to1 liter
4x Resolving-gel buffer1.5M Tris-Cl (pH 8.8)
8x Stacking-gel buffer1M Tris-Cl (pH 6.8)
1x SDS gel-loading buffer50mM Tris-Cl (pH 6.8)
100mM dithiothreitol
2% SDS
0.1% bromophenol blue
10% glycerol



RESOLVING GEL MIXTURE
Resolving Gel (30ml)6%8%10%12%15%
H2O16.214.212.210.27.2
30% acrylamide mix6.08.0101215
4x resolving-gel buffer7.57.57.57.57.5
20% SDS0.150.150.150.150.15
20% APS0.150.150.150.150.15
TEMED0.0240.0180.0120.0120.012

Resolving Gel (10ml)6%8%10%12%15%
H2O5.44.74.13.42.4
30% acrylamide mix2.02.73.345
4x resolving-gel buffer2.52.52.52.52.5
20% SDS0.050.050.050.050.05
20% APS0.050.050.050.050.05
TEMED0.0080.0060.0040.0040.004



STACKING GEL MIXTURE
Stacking Gel (10ml)
H2O6.9 ml
30% acrylamide mix1.7 ml
8x stacking-gel buffer1.25 ml
20% SDS0.05 ml
20% APS0.05 ml
TEMED0.01 ml



SOLUTIONS FOR STAINING & DESTAINING
Staining solutionmethanol90 ml
H2O90 ml
acetic acid20 ml
Coomassie Brilliant Blue R2500.25 g
Dissolve well and filtrate through a filter paper.
Destaining solutionisopropanol12.5%
glacial acetic acid10%
Addition of small pieces of used X-ray film with the
gel will accelerate the destaining process.



SOLUTION FOR DRYING

20% methanol, 3% glycerol in diltilled water




siraisi@kuchem.kyoto-u.ac.jp

To index

To Shiraishi's Home Page
To Biochmistry Lab. Home Page