Abstract
We report a streamlined procedure to efficiently carry samples from chromatin to qPCR-compatible DNA in as little as 4 h. We use this streamlined ChIP to quantify histone H3 modifications at active (cad) and repressed (T early alpha) promoters in a Rag1-deficient pro-T cell line after 1-2 h IP. We further show that the protocol readily quantified histone modifications in chromatin from 10(4) Rag-deficient DN thymocytes. Taken together, these data outline a simple, cost-effective procedure for efficient ChIP analysis.
Publication types
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Research Support, N.I.H., Extramural
MeSH terms
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Animals
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Aspartate Carbamoyltransferase / genetics
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Aspartate Carbamoyltransferase / metabolism
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Carbamoyl-Phosphate Synthase (Glutamine-Hydrolyzing) / genetics
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Carbamoyl-Phosphate Synthase (Glutamine-Hydrolyzing) / metabolism
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Cell Line
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Chromatin / genetics
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Chromatin / isolation & purification*
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Chromatin / metabolism
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Chromatin Immunoprecipitation / methods*
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DNA-Binding Proteins / genetics
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DNA-Binding Proteins / metabolism
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Dihydroorotase / genetics
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Dihydroorotase / metabolism
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Gene Knockdown Techniques
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Histones / genetics
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Histones / metabolism*
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Mice
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Mice, Knockout
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RNA Polymerase II / genetics
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RNA Polymerase II / metabolism
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T-Lymphocytes / metabolism
Substances
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CAD trifunctional enzyme
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Chromatin
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DNA-Binding Proteins
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Histones
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Rag2 protein, mouse
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Aspartate Carbamoyltransferase
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RNA Polymerase II
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Dihydroorotase
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Carbamoyl-Phosphate Synthase (Glutamine-Hydrolyzing)