C.B.S. Scientific SGU-2640T-02 - Manual (Page 12)

C.B.S. œ Scientific Submarine/Horizontal 12
SECTION 4
Running Conditions
4.1 Recommended Power
As a rule, optimal resolution of larger molecules is achieved during longer runs at lower voltages,
whereas smaller molecules require shorter runs at higher voltages. Applied voltage gradients
can therefore be anywhere in the range of 1-10 VDC/cm of gel. Using the standard buffer
systems listed below, most runs will use 5 VDC/cm as a general rule.
The usual run time will vary for the voltage chosen but should range from 15 to 60 minutes.
Nucleic acid migration is monitored by the progress of marker dyes. Constant power is not a
necessity, but it produces uniform heat throughout the run, therefore minimizing band diffusion.
Be sure the polarity is correct i.e. that the DNA is loaded near the cathode (black electrode) to
run toward the anode (red terminal).
Agarose gels may be stored for several days at 4
°C wrapped in plastic wrap. Seakem Agarose
(FMC) is used (normally) for preparative and analytical gels. Other types of agarose can be used
for special purposes.
4.2 Recommended Buffer and Agarose Volumes
Cat. # Gel Bed dimensions Buffer
(mls) Agarose
for 1cm
thick slab
(mls)
SGU-014T-02 14cm x 20cm 1500 290
SGU-020T-02 20cm x 20cm 1850 400
SGU-030T-02 20cm x 30cm 1950 600
SGU-040T-02 20cm x 40cm 2050 800
SGU-2614T-02 26cm x 14cm 2200 400
SGU-2626T-02 26cm x 26cm 2600 700
SGU-2640T-02 26cm x 40cm 3400 1100
4.3 Recommended Buffers and Solutions*
Type Concentrated Stock/liter Final Concentration
TAE 50X - 242 gm Tris base 1X - 0.04M Tris-acetate
(Tris-acetate) 57.1ml glacial acetic acid 0.001M EDTA
100 ml 0.5M EDTA (pH 8.0)
TBE 5X - 54 gm Tris base 0.5X - 0.045M Tris-borate
(Tris-borate) 27.5 gm boric acid 0.001M EDTA
20 ml 0.5M EDTA (pH 8.0)
10X - Loading Buffer (DNA)*
0.25% Bromophenol blue
0.25% Xylene cyanol
20% Ficoll Type 400
0.1M EDTA, pH 8.0
Ethidium Bromide Staining
EtBr can be premixed with buffers and agarose for use during electrophoresis. Add to
agarose only after temperature has fallen below 55
°C. Gels can also be stained after
electrophoresis in a soaking tray. Use EtBr at a final concentration of 0.1
μg/ml from a
1mg/ml stock solution. Ethidium Bromide is a powerful mutagen. ALWAYS wear gloves,
eye protection and protective clothing. Dispose of solutions in accordance with the safety
regulations of your institution.