Owner manual

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Cipher DGGE Instructions 2/28/14
SECTION 6
Recommended Buffers and Reagents
Warning: Do not mix buffer in the DGGE tank!
6.1 Stock Solutions
Acrylamide Stock Solution 40% Acrylamide/Bis (37.5:1)
For 100ml use the following:
38.93 g of acrylamide
1.07 g of Bis-acrylamide
Add dH
2
O (or RO-H
2
O) to 100ml. Do not autoclave
50x TAE Gel Running Buffer
For 1 liter use the following:
242g Tris Base
57.1ml Glacial acetic acid
100ml .5 M EDTA, pH 8.0
Add dH
2
O (or RO-H
2
O) to 1 liter
Denaturant Stock Solution (7.5% gel)
For 100ml use:
Add dH
2
0 (or RO-H
2
O) to 100ml
Store denaturant stock solutions at 4º C in amber bottles
Polymerization Catalysts:
1. TEMED (10ul/ml gel solution)
2. APS Stock Solution (.6ul/ml gel solution)
For 50 ml of APS stock (10%) use 5 g of ammonium persulphate and water to
50ml. Store at 4ºC.
Neutral Loading Solution
For preparation of 10ml:
20% Ficoll or sucrose, 2 g of Ficoll 400 or sucrose
10mM Tris-HCl, pH 7.8, 100ul of 1.0 M Tris-HCl, pH 7.8
1mM EDTA, 20 ul of 0.5 M EDTA, pH 8.0
0.1% dye (BPB), 10 mg of dye
Water to 10ml
Percentage 10 20 30 40 50 60 70 80 90 100
40% Acrylamide/
Bis (ml)
18.8 18.8 18.8 18.8 18.8 18.8 18.8 18.8 18.8 18.8
50% TAE Buffer
(ml)
2 2 2 2 2 2 2 2 2 2
Formamide (ml) 4 8 12 16 20 24 28 32 36 40
Urea (g) 4.2 8.4 12.6 16.8 21 25.2 29.4 33.6 37.8 42
SECTION 7
Maximum Well/Comb Volumes
Vertical Comb Options, deeper wells, overall length = 13.1cm
Cat. # Tooth Depth
(mm)
# of
Teeth
Thickness
of Teeth
Width of
Teeth
Recommended
Max. Sample
Vol./Well (µl)*
VGC-0716 19.1 16 0.75 5.2 55
VGC-0720 17.7 20 0.75 4.2 42
VGC-0722 17.7 22 0.75 4.35 25
VGC-0729 17.7 29 0.75 2.9 29
VGC-0730 17.7 30 0.75 2.8 28
VGC-1016 19.1 16 1.0 5.2 74
VGC-1020 19.1 20 1.0 4.2 55
VGC-1022 17.7 22 1.0 4.35 57
VGC-1029 17.7 29 1.0 2.9 38
VGC-1030 17.7 30 1.0 2.8 37
VGC-1516 19.1 16 1.5 5.2 110
VGC-1520 17.7 20 1.5 4.2 83
VGC-1522 17.7 22 1.5 4.35 85
VGC-1529 17.7 29 1.5 2.9 58
VGC-1530 17.7 30 1.5 2.8 56
Vertical Comb Options, short wells, overall length = 13.1cm
Cat. # Tooth depth
(mm)
# of
Teeth
Thickness
of Teeth
Width of
Teeth
Recommended
Max. Sample
Vol./Well (µl)*
VGC-0716-177 10 16 0.75 5.2 29
VGC-0720-177 10 20 0.75 4.2 23
VGC-0722-177 10 22 0.75 4.35 25
VGC-0729-177 10 29 0.75 2.9 16
VGC-0730-177 10 30 0.75 2.8 16
VGC-1016-177 10 16 1.0 5.2 39
VGC-1020-177 10 20 1.0 4.2 32
VGC-1022-177 10 22 1.0 4.35 32
VGC-1029-177 10 29 1.0 2.9 22
VGC-1030-177 10 30 1.0 2.8 21
VGC-1516-177 10 16 1.5 5.2 58
VGC-1520-177 10 20 1.5 4.2 47
VGC-1522-177 10 22 1.5 4.35 49
VGC-1529-177 10 29 1.5 2.9 33
VGC-1530-177 10 30 1.5 2.8 32
* Maximum loading volume is calculated by taking 75% of the total tooth volume.