The fellows just met in Madrid on the week of the 18th-22nd of September to learn how to perform a ChIP-seq on Brassica napus. They first went in-depth into the theory and then they went hands-on the protocol.

But… What is ChIP-seq?

ChIP-seq is a tool to investigate the epigenomes of multiple cell types and their underlying mechanisms. It is based on chromatin immunoprecipitation (ChIP) which is a protein-DNA binding in vivo assay technique. The ChIP antibodies are specific to our protein or nucleosome of interest. ChIP is followed by sequencing (the –seq part of ChIP-seq), which allows to identify and sequence our enriched DNA fragments (Park, 2009).

There are four basic steps for ChIP, as you can see in the image below. Firstly, crosslinking is the way to improve the connection between protein and chromatin to preserve the interaction for further antibody targeting. There are two common methods to do crosslinking, using formaldehyde or ultraviolet (UV) light. For this ChIP-seq course, we used 1% formaldehyde. After crosslinking, we need to extract chromatin, and shearing/sonicate them. Shearing step to solubilize the chromatin which is very important to get enough chromatin yield. In addition, a proper shearing strategy to generate desirable DNA size (300bp-500bp) is good for immunoprecipitation and signal detection. The next step is to remove these fragments without the connection between the targeted protein and chromatin. To achieve this, we need to use antibodies to bind with targeted proteins where the antibody can stick to magnetic beads. So, the non-target proteins can be washed out, followed by eluting the targeted protein from the beads. Then, de-crosslinking means removing the proteins from the DNA to make the purified DNA accessible easily. Lastly, indexing barcodes will be added to the purified DNA for individual sample identification. If the quality control test shows positive results, the library could be used for ChIP-seq.


(Daniel et al., 2014)

From this we understand how ChIP sequencing is carried out and the steps involved in successful preparation of a library for sequencing, further using various sequencing platforms sequencing of the prepared libraries can be carried out.

With excellent guidance by Dr. Pedro Crevillén and Dr. Jose A. Abelenda, fellows of the EpiSeedLink doctoral network were skillfully trained in ChIP sequencing. This will immensely help each of the fellows in the next phase of their PhD journey, where they will incorporate the knowledge gained in practical terms.

Each fellow will use ChIP sequencing technique to uniquely benefit their research project, resulting in generating enormous data that will help the research community in understanding the dynamics of both Arabidopsis and Brassica napus genome.


Text by Shreyas Padmanabha Sharma Beedubail, PhD Student EpiSeedLink Marie Skłodowska-Curie Actions



  1. Daniel, B., Balint, B. L., Nagy, Z. S., & Nagy, L. (2014). Mapping the genomic binding sites of the activated retinoid X receptor in murine bone marrow-derived macrophages using chromatin immunoprecipitation sequencing. Steroid Receptors: Methods and Protocols, 15-24.
  2. Park, P. J. (2009). ChIP–seq: advantages and challenges of a maturing technology. Nature reviews genetics10(10), 669-680.



Frontal image by <a href=””>Freepik</a>