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Fig. 1.

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Fig. 1. Schematic representation of assays for studying chromosome architecture. For 3C-based methods, nuclei are first treated with appropriate fixatives (i.e., formaldehyde, DSG, etc.). In GAM, cryosections are cut from paraformaldehydefixed and sucrose-embedded samples. In 3C, 4C and 5C, fixed nuclei are treated with restriction enzymes, ligated, and the ligation frequency is measured by PCR or NGS. For Hi-C, Plac-seq, and cHi-C, chromosomal DNA is digested by restriction enzymes while micro-C uses MNase for finer resolution. The digested DNA ends are repaired with biotin-labeled nucleotides followed by blunt-end ligation. The ligated biotin-labeled contacts are sheared and purified with streptavidin beads prior to NGS sequencing. In Plac-seq and cHi-C, antibody pull-down or RNA oligo-mediated DNA pull-down is performed for target enrichment, respectively. In the SPRITE method, crosslinked chromatin is fragmented by sonication, each interacting complex is uniquely tagged by multiple rounds of split-pool barcoding, and the final material is sequenced. In the GAM method, the DNA contents from cryosections are extracted, fragmented, and sequenced. Appropriate computational analysis of the sequencing data from each approach is necessary to detect physical interactions between genomic loci.
J. Microbiol. Biotechnol. 2022;32:1515~1526
© J. Microbiol. Biotechnol.