If 4C and 5C are sequels to 3C, then Hi-C is a total franchise reboot (and as The Dark Knight showed us, that can be a great thing). Hi-C is the first of the 3C derivative technologies to be truly genome-wide.
After the restriction digest, biotin-labeled nucleotides are annealed to the fragment end overhangs. The blunt ends of the DNA fragments are ligated. A biotin pull-down then isolates only ligated fragments. The library is then sequenced and mapped to the genome allowing fragment identity and abundance to be detected. While Hi-C is high-resolution compared to 3C, it is still not as high as some 4C variants (de Wit and de Laat, 2012; Dekker, 2006).
This means that experiments examining the broad picture of interactions across the genome use Hi-C, while high-resolution interactions of a specific site use 4C. Another disadvantage of Hi-C is workload increase compared to 4C or 5C: since the resolution increases, the sequencing depth must also increase (Dekker, 2006). Hi-C was first described in 2009 where it was used to find a new level of genomic organization of human chromatin domains (Lieberman-Aiden et al., 2009). Since, Hi-C has become the method of choice for researchers looking to characterize the structure of the entre genome, especially when lacking a priori knowledge about that to expect (see Jin et al., 2013).
For additional reading about chromosome conformation capture methods, check out this article on Hi-C and related methods from our friends at Active Motif.
Hi-C Additional Reading
Jin, F., Li, Y., Dixon, J.R., Selvaraj, S., Ye, Z., Lee, A.Y., Yen, C.A., Schmitt, A.D., Espinoza, C.A., and Ren, B. (2013). A high-resolution map of the three-dimensional chromatin interactome in human cells. Nature 503, 290-294.
This paper uses Hi-C to map the structure of human chromosomes. The authors propose two general principles of chromatin organization from their data. First, that enhancer interaction loops exist long before actual induction occurs. Also, chromatin structure appears to be stable each cell type, influencing gene-expression patterns.
This paper describes Hi-C in detail from start to finish. It is both an excellent introduction to the technology, as well as a good technical resource.
- de Wit, E., and de Laat, W. (2012). A decade of 3C technologies: insights into nuclear organization. Genes Dev. 26, 11-24.
- Dekker, J. (2006). The three ‘C’ s of chromosome conformation capture: controls, controls, controls. Nat. Methods 3, 17-21.
- Lieberman-Aiden, E., van Berkum, N.L., Williams, L., Imakaev, M., Ragoczy, T., Telling, A., Amit, I., Lajoie, B.R., Sabo, P.J., Dorschner, M.O., et al. (2009). Comprehensive mapping of long-range interactions reveals folding principles of the human genome. Science 326, 289-293.