Solutions for Txn Lab
FIR 2007
NUCLEAR EXTRACT ISOLATION
Solution Final Conc Stock soln concentration Vol (ul) for 10 ml Final
HEGD (hypotonic buffer) 25 mM 1M HEPES pH 7.9 250
(make FRESH) 1 mM 1M DTT 10
1.5mM 0.5M EDTA pH 8.0 30
10% 50 % glycerol 2000
1X *50X complete protease inhibitor mix 200
H2O 7510
Vol (ul) for 2ml Final
HEGDK (extraction buffer) 25 mM 1M HEPES pH 7.9 50
(make FRESH) 1 mM 1M DTT 2
1.5mM 0.5M EDTA pH 8.0 6
10% 50% glycerol 400
0.5M 1.0M KCl 1000
1X *50X complete protease inhibitor mix 40
H2O 502
For 0.5 ml Final
8-Bromo-cAMP Stock Soln. 100mM 8-Bromo- 100mM 8-Bromo-cAMP Disolve 17.20 mg 8-bromo-
Protien Assay Solutions
Vol for 1ml 2X Buffer
2X CREB EMSA Buffer 10mM Tris (7.4) 1M Tris (7.4) 20
50mM NaCl 5M NaCl 20
1mM MgCl2 1M MgCl2 2
0.5mM EDTA 0.5M EDTA 2
5% Glycerol 50% Glycerol 200
0.5mM DTT 1M DTT 1
125ug/ml BSA 20mg/ml BSA 12.5
50 ug/ml dIdC 0.5mg/ml 200
H2O 462.5
Solutions for Txn Lab
FIR 2007
Vol (ul) for 1ml Final
ChIP Lysis Buffer 10mM 0.5M EDTA 20
50mM 1M Tris pH 8.1 50
1% 10% SDS 100
1X *50X complete protease inhibitor mix 20
H2O 790
Vol (ul) for 4ml Final
ChIP Dilution Buffer 16.7mM 1M Tris pH 8.1 67
167mM 5M NaCl 134
1.22mM 0.5M EDTA 9.8
1.10% 10% Triton X-100 440
0.01% 10% SDS 4
1X *50X complete protease inhibitor mix 80
H2O 3266
Vol (ul) for 4ml Final
ChIP Low-Salt Wash Buffer 20mM 1M Tris pH 8.1 80
150mM 5M NaCl 120
2mM 0.5M EDTA 16
1% 10%Triton X-100 400
0.10% 10% SDS 40
1X *50X complete protease inhibitor mix 80
H2O 3264
Vol (ul) for 4ml Final
ChIP High-Salt Wash Buffer 20mM 1M Tris pH 8.1 80
500mM 5M NaCl 400
2mM 0.5M EDTA 16
1% 10% Triton X-100 400
0.10% 10% SDS 40
1X *50X complete protease inhibitor mix 80
H2O 2984
Vol (ul) for 4ml Final
ChIP LiCl Wash Buffer 250mM 5M LiCl 200
1% 10% NP-40 400
1% 10% Na Deoxycholate 400
10mM 1M Tris pH 8.1 40
1mM 0.5M EDTA 8
1X *50X complete protease inhibitor mix 80
H2O 2872
Vol (ul) for 8ml Final
ChIP TE 10mM Tris (8.0) 1M Tris (8.0) 80
1mM EDTA 0.5M EDTA 16
1X *50X complete protease inhibitor mix 160
H2O 7744
Vol (ul) for 2ml Final
ChIP Elution Buffer 100mM 1M NaHCO3 200
1% 10% SDS 200
H2O 1600
Vol/rxn
PCR Cocktail H2O 14
1X 10X PCR Buffer 2.5
20 mM MgCl2 50 mM MgCl2 1
0.25 mM dNTPs 25 mM dNTP's 0.25
0.25 uM *For [6.25 pmol/ul] 1
0.25 uM *Rev [6.25 pmol/ul] 1
Platinum Taq DNA Polymerase 0.25
DNA Template 5
For 0.2 ml Final
Roche protease inhibitor stock 50X Stock Soln. 50X Disolve 1 complete mini tablet
into 200 ul sterile H2O
* Protease inhibitors must be added fresh to solutions. If you have extra protease inhibitor, store it at
and use it for solutions made at a later time.
Solutions for Txn Lab
FIR 2007
Solutions for Txn Lab
FIR 2007
For 1 ml Final
Disolve 43.01 mg 8-
Solutions for Txn Lab
FIR 2007
Vol (ul) for 4ml Final
80
200
400
80
3160
Vol (ul) for 15ml Final
251
502
37
1.65 ml
15
300
12.24 ml
Vol (ul) for 10ml Final
200
300
40
1 ml
100
200
8.16 ml
Vol (ul) for 10ml Final
200
1 ml
40
1 ml
100
200
7.46 ml
Vol (ul) for 10ml Final
500
1 ml
1 ml
100
20
200
7.18 ml
Vol (ul) for Xml Final
Vol (ul) for Xml Final
Vol/15 rxn
210
37.5
15
3.75
15
15
3.75
For 1 ml Final
Dissolve 1 complete or
5 complete mini tablets
into 1ml sterile H2O
bitor, store it at -20oC
Solutions for Txn Lab
FIR 2007
Reaction Protein Concentration Sample Vol. (ul) Assay Conc.
Sample (ug/ul) (ug/ml)
Blank None 0 0 0
1 BSA 0.5 1 2
2 BSA 0.5 2 4
3 BSA 0.5 3 6
4 BSA 0.5 4 8
5 BSA 0.5 5 10
6 BSA 0.5 6 12
7 BSA 0.5 10 20
8 BSA 0.5 15 30
9 MA-10 NE ? 1 ?
10 MA-10 NE ? 2 ?
11 MA-10 NE ? 3 ?
H20 Vol. (ul) Dye Conc. Vol.
(ul)
800 200
799 200
798 200
797 200
796 200
795 200
794 200
790 200
785 200
799 200
798 200
797 200