Gel protocol

Protocols to make SDS-PAGE gel
Resolving Gel
                                         12%(10ml)                     14%(10ml)
                    
30%  Acryl-Bis:            4.0ml                                 3.5ml
1.5M Tris/HCl, pH=8 : 2.5ml                                 2.5ml
10%SDS                         100ul                                 100ul
Distilled water                3.3ml                                3.8ml
10%APS                         100ul                                 100ul
TEMED                           5ul                                      5ul

Stacking Gel                    4%(10ml)
30% Acryl-Bis                 1ml
0.5M Tris/HCl pH=6.8    2.5ml
10%SDS                           100ml
Distilled water                  6ml
10%APS                            100ul
TEMED                              10ul
Running Buffer
10x
Tris                    30g/L
Glycine               144g/L
SDS                   10g/L
To make the running buffer, the 10X is diluted 10times, the 10X buffer is made for convinience.
4X sample buffer
Tris, pH=8, 250mM
Glycerol     40%(v/v)
SDS(powder)   9.2%(w/v)
Beta-Mercaptoethanol 20%(v/v)
Bromophenol blue    Trace of 2mL of a solution of 0.1%(Bromophenol is insolutable in water)
Staining solution
1 tablet of Coomassie in 1.0L of 0.5%(v/v) acetic acid.
De-staining solution
30% acetic acid
When casting gel some notices need to be taken in to your mind
- check the wells whether there is a leaking
- running voltage: 100-200 usually 100 or 90 V