Mirmira Lab ChIP Protocol
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Reagents:
ChIP sonication Buffer (1% Triton X-100, 0.1% Deoxycholate, 50 mM Tris 8.1, 150 mM NaCl, 5 mM EDTA):
10 ml 10% Triton X-100
1 ml 10% Deoxycholate
5 ml 1 M Tris-Cl pH 8.1
1 ml 0.5 M EDTA
3 ml 5 M NaCl
80 ml Water
Just before use, add 10 ul Aprotinin, 10 ul Leupeptin, and 5 ul PMSF to each 10 ml.
High Salt Wash Buffer (1% Triton X-100, 0.1% Deoxycholate, 50 mM Tris-8.1, 500 mM NaCl, 5 mM EDTA)
10 ml 10% Triton X-100
1 ml 10% Deoxycholate
5 ml 1M Tris-8.1
1 ml 0.5M EDTA
10 ml 5M NaCl
73 ml Water
(LiCl Immune Complex Wash Buffer)
25 ml 1M LiCl
5 ml 10% IGEPAL
5 ml 10% Deoxycholate
1 ml 1M Tris-8.1
200 ul 0.5M EDTA
64 ml Water
Protease inhibitors (add 10 ul of each to 10 ml of PBS or sonication buffer)
Leupeptin 2 mg/ml in water
Aprotinin 2 mg/ml in water
PMSF 0.2 M
1X TE Buffer (10mM Tris, 8.1, 1 mM EDTA)
0.5 M EDTA
Protein A/G Agarose (Santa Cruz #sc-2003)
Proteinase K (19 mg/ml, Boehringer Mannheim # 1964372)
10X proteinase K buffer
Elution Buffer (1% SDS, 0.1 M NaHCO3, 0.01 mg/ml Herring sperm DNA)
10 mg/ml Herring Sperm DNA
37% Formaldehyde (ACS reagent grade)
1.25 M glycine
