Chromatin Immunoprecipitation Workshops

The next ChIP workshop has not yet been scheduled.  If you would like to participate, please contact Charlie or Heather in order to be notified right away.  You can download a copy of the last announcement to get some information about the structure of the workshop.  The workshop begins with a presentation on the background of ChIP-chip and in-depth information about some of the manipulations we carry out.  A copy of the latest Powerpoint presentation from October 21st, 2009 is available for download.  At the end of the workshop, Dr. Peggy Farnham presents on the uses of ChIP and ChIP-chip, then takes questions and leads a discussion about the technical details of the procedure.  These discussions are open to the community.  Download Dr. Farnham's latest presentation from octoboer 23rd, 2009. 

Representative PCR Results from July 27th, 2007 Workshop:

Upper gel group 1, Lower gel group 3

                        Sample                              Amplified Fragment

lane 1    anti-pol2 precipitated       pol2 promoter (positive control)
lane 2    IgG precipitated                pol2 promoter (positive control)
lane 3    Input DNA, 1:20 dilution  pol2 promoter (positive control)
lane 4    Input DNA, 1:5 dilution    pol2 promoter (positive control)
lane 5    anti-pol2 precipitated       DHFR 3' UTR (negative control)
lane 6    IgG precipitated                DHFR 3' UTR (negative control)
lane 7    Input DNA, 1:20 dilution  DHFR 3' UTR (negative control)
lane 8    Input DNA, 1:5 dilution    DHFR 3' UTR (negative control)
lane 9    MW marker

 

Both groups saw the expected results:

1. Strong amplification of the pol2 promoter in the pol2 precipitated samples compared to the IgG samples
2. No difference in amplification of the DHFR 3' UTR between the pol2 and IgG precipitated samples
3. Much higher signal ratio of pol2 precipitated:input DNA in the positive control compared to the negative control.

For a more detailed explanation of how to interpret a ChIP experiment, download this explanatory document.

For information on calculating the efficiency of your precipitation, download this diagram.

Chromatin Immunoprecipitation Resources

ChIP-chip Tech Guide. A recent tech guide from Genome Web that provides a nice overview of some basic ChIP and ChIP-ship issues from experts in the field. It's a bit slick with a few ads but it's up to date and free.

A good resource for information about primary antibodies that have been shown to work in chromatin immunoprecipitation is the chIP on chip.org site hosted by Bing Ren's lab at UCSD.  Researchers can scan a number of commercial antibodies to see if they've been deemed chIP-worthy.  The site is a little out of date but it still has some useful information.

You might also be interested in looking over a few papers that provide background on the ChIP-chip procedure and illustrate some applications of the technology.

High Throughput Screening of Chromatin Immunoprecipitates Using CpG-Island Microarrays Methods in Enzymology article explaining the basic protocol.  While technical developments have advanced, this paper provides useful background information and rationales for the procedures used.

Use of Chromatin Immunoprecipitation to Clone Novel E2F Target Promoters Primary publication using a protocol very similar to the one used in the workshop.

Locating Mammalian Transcription Factor Binding Sites: A Survey of Computational and Experimental Techniques Recent review outlining the application of ChIP-chip plus in silico techniques for genome wide sequence element characterizations.

Chromatin Immunoprecipitation and Microarray-Based Analysis of Protein Location  Recent techniques oriented paper from another group describing entire ChIP-chip procedure, with detailed protocols and helpful troubleshooting information.

Two-step Cross-linking Method for Identification of NF-kB Gene Network by Chromatin Immunoprecipitation  Demonstration and theoretical discussions on the use of additional cross linkers besides formaldehyde in order to expand information obtained in the ChIP assay.

In vivo Dual Cross-linking for Identification of Indirect DNA-associated Proteins by Chromatin Immunoprecipitation Similar to above paper, explores crosslinkers with different spacer lengths to identify additional proteins in transcription complexes.

Comparison of Sample Preparation Methods for ChIP-chip Assays Recent collaborative effort from the Farnham group and core facility on the use of whole genome amplification (WGA) reagents for ChIP sample amplification.

WGA Publication Supplemental Information Protocols accompanying above paper, includes comprehensive ChIP protocol and WGA protocol.

Whole Genome Amplification Protocol WGA protocol alone, update March 27, 2007.

Chromatin Immunoprecipitation Protocol Latest version of the ChIP protocol, updated April 9th, 2008.

Workshop Chromatin Immunoprecipitation Protocol Nearly identical to the above protocol, more configured towards workshop specific manipulations (updated April 9th, 2008).

Chromatin Immunoprecipitation Supplies Vendor and order information for the supplies we use for ChIP experiments (updated April 9th, 2008).

Farnham Lab Web Site Follow the links from this page to more protocols for ChIP and ChIP-chip, primary publications utilizing these techniques, software analysis tools, and links to numerous other related resources.

Epigenome Network Web Site.  European Union based web site containing solicited protocols from international experts in the fields of ChIP, ChIP-chip, and other epigentic analytical techniques. Our WGA protocol is on this site.

Micro-ChIP protocol.  Protocol developed in Farnham lab for performing ChIP on very small cell numbers, as few as 10,000 cells.

The Chromatin Remodeler SPLAYED Regulates Specific Stress Signaling Pathways.  Publication from Walley et al., Dehesh lab UC Davis, using ChIP techniques learned in the workshop.