Imprint® Chromatin Immunoprecipitation Kit

Protocol tips

Upstream tips	Protocol tips	Downstream tips
-If any reagent has formed a precipitate, warm at 55 °C until dissolved. Allow to equilibrate to room temperature before use.	-Use 1 µg of antibody per well. -The range of cells that can be used with this kit is 0.2–2.5 x 105/well/ChIP sample. Using more than 0.5 million cells per well will increase the non-specific binding and reduce the yield and specificity of the ChIP reaction. If higher yield of ChIP DNA is desired for downstream applications (e.g. ChIP-Seq., ChIP-chip): a) DNA could be pooled by eluting consecutively (with 50 µl of elution buffer) from multiple individual ChIP reactions.

Upstream tips
-If any reagent has formed a precipitate, warm at 55 °C until dissolved. Allow to equilibrate to room temperature before use.
Protocol tips
-Use 1 µg of antibody per well. -The range of cells that can be used with this kit is 0.2–2.5 x 105/well/ChIP sample. Using more than 0.5 million cells per well will increase the non-specific binding and reduce the yield and specificity of the ChIP reaction. If higher yield of ChIP DNA is desired for downstream applications (e.g. ChIP-Seq., ChIP-chip): a) DNA could be pooled by eluting consecutively (with 50 µl of elution buffer) from multiple individual ChIP reactions.

Publication protocol

ChIP assays were performed using the Imprint Ultra Chromatin Immunoprecipitation Kit (Sigma, St. Louis, Missouri). Both quantitative PCR and standard agarose gel-based PCR were used to monitor the ChIP results. PCRs contained 1.25 U of Taq polymerase enzyme (Promega Corp., Madison, Wisconsin), 1× reaction buffer, 1.5 mM MgCl2, 0.5 mM deoxynucleotide triphosphate (Invitrogen), 357 nM primers, and 2 μL of DNA from the ChIP assay in a final volume of 50 μL. PCR products (20 μl) were loaded on 1.2% agarose gels, and electrophoresis was performed at 100 V for 1.5 hours in 1× TAE buffer.

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Manufacturer protocol

Download the product protocol from Sigma-Aldrich for Imprint® Chromatin Immunoprecipitation Kit below.

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