ChIP Mouse - Osteoblasts

DNA-protein interactions are studied by using ChIP. The basic steps in this technique are crosslinking, sonication, immunoprecipitation, and analysis of the immunoprecipitated DNA. During ChIP, if chromatin is under-fragmented or fragments are too large which can lead to the increased background and lower resolution. Shorter cross-linking times (5-10 min) and/or lower formaldehyde concentrations (<1%) may improve shearing efficiency. If Chromatin is over-fragmented, then optimize shearing conditions for each cell type to improve ChIP efficiency. Over-sonication of chromatin may disrupt chromatin integrity and denature antibody epitopes. If you do not see any product or very little product in the input PCR reactions, add 5–10 μg chromatin per IP.

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Found 2 matching solutions for this experiment

ChromaFlash Chromatin Extraction Kit

Epigentek

Upstream tips	Protocol tips	Downstream tips
- Spin the solution down to the bottom prior to use. - Check if any buffers contain salt precipitates before use. If so, shake the buffer until the salts are re-dissolved.	-For optimal preparation, the input amount should be 1 to 5 x 106 cells or 50 to 200 mg tissues. -Avoid multiple freeze/thaw cycles.

Upstream tips
- Spin the solution down to the bottom prior to use. - Check if any buffers contain salt precipitates before use. If so, shake the buffer until the salts are re-dissolved.
Protocol tips
-For optimal preparation, the input amount should be 1 to 5 x 106 cells or 50 to 200 mg tissues. -Avoid multiple freeze/thaw cycles.

Manufacturer protocol Publication protocol 1 Relevant paper

truChIP Ultra-Low Chromatin Shearing Kit with Formaldehyde

Covaris

Upstream tips	Protocol tips	Downstream tips
	24 h after silencing, the cells were washed with PBS and complete MEM media was added to each well and left for additional three days, after which ChIP was performed using truChIP™ Ultra Low Cell Chromatin Shearing Kit (Covaris, Inc., Woburn, Massachusetts).

Protocol tips
24 h after silencing, the cells were washed with PBS and complete MEM media was added to each well and left for additional three days, after which ChIP was performed using truChIP™ Ultra Low Cell Chromatin Shearing Kit (Covaris, Inc., Woburn, Massachusetts).

Manufacturer protocol Publication protocol 1 Relevant paper

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