4DN Protocols

Collection of genomic technologies currently in use or being developed in the 4DN network.

ProtocolDate of approvalDescription
iMARGIAugust 18, 2020
in situ Mapping of RNA-chromatin interactions (iMARGI) protocol for mapping of chromatin-associated RNAs.
August 18, 2020
PLAC-seq/HiChIP/in situ ChIA-PET protocols to detect and quantify chromatin contacts anchored at genomic regions associated with specific DNA binding proteins or histone modifications.
Single-cell Hi-C ProtocolApril 17, 2018Single-cell Hi-C Protocol and Quality Control Standards
DNase Hi-C protocolJanuary 16, 2018Mapping 3D genome architecture through in situ DNase Hi-C
DamID seq protocolDecember 12, 2017
The Official 4DN Standard DAM-ID seq Protocol from the van Steensel Lab.
ChIA-PET data analysisOctober 17, 2017
ChIA-PET data processing pipeline and standards, 4D Nucleome Consortium
ChIA-PET experimental protocolOctober 17, 2017ChIA-PET Protocol and Standards, 4D Nucleome Consortium
E/L Repli-seqJune 20, 2017
E/L Repli-seq: Protocol and Quality Control Standards, 4D Nucleome Consortium
E/L Repli-seqJune 20, 2017
4D Nucleome Consortium, Overall Standards and Guidelines for E/L Repli-seq Experiments
Hi-CFebruary 21, 2017Standards and Guidelines for Hi-C experiments
In situ Hi-CFebruary 21, 2017Protocol and Quality Control Standards for in situ Hi-C experiments

Other relevant protocols

Technology (Protocol)Reference paperDescription
3C-seqChromosome conformation capture techniques are used to analyze the organization of chromatin in a cell by quantifying the interactions between genomic loci that are nearby in 3-D space. 3C quantifies interactions between a single pair of genomic loci (one-vs-one).
4C-seqChromosome conformation capture-on-chip (4C) captures interactions between one locus and all other genomic loci (one-vs-all).
5C-seqChromosome conformation capture carbon copy (5C) detects interactions between all restriction fragments within a given region, with this region's size typically no greater than a megabase (many-vs-many).
Capture Hi-CCapture Hi-C (CHi-C) is is an adapted technology that selects and enriches few hundred promoters for genome-wide, long-range contacts of both active and inactive promoters.
ChIA-PETChromatin Interaction Analysis by Paired-End Tag Sequencing (ChIA-PET) is a technique that incorporates chromatin immunoprecipitation (ChIP)-based enrichment, chromatin proximity ligation, Paired-End Tags, and High-throughput sequencing to determine de novo long-range chromatin interactions genome-wide.
COLACOLA (Concatamer Ligation Assay) is a modified in situ Hi-C protocol that uses CviJI restriction enzyme to digests chromatin into much finer fragments the original Hi-C method, in order to increase the proportion of reads containing three or more nearby fragments.
CUT&RUNCUT&RUN is an efficient targeted nuclease strategy for high-resolution mapping of DNA binding sites. Cleavage Under Targets and Release Using Nuclease (CUT&RUN) is an antibody-targeted chromatin profiling method in which micrococcal nuclease tethered to protein A binds to an antibody of choice and cuts immediately adjacent DNA, releasing DNA bound to the antibody target. The procedure is carried out in situ and produces precise transcription factor or histone modification profiles while avoiding crosslinking and solubilization issues. Extremely low backgrounds make profiling possible with typically one tenth of the sequencing depth required for ChIP, and permit profiling using low cell numbers without loss of quality. CUT&RUN can also be used to map long-range genomic contacts.
DamIDDamID enables mapping genome-wide occupancy of interaction sites in vivo, based on the expression of a fusion protein consisting of a protein of interest and DNA adenine methyltransferase (Dam). This leads to methylation of adenines near sites where the protein of interest interacts with the DNA. These methylated sequences are subsequently amplified by a methylation-specific PCR protocol and identified by hybridization to microarrays.
Dnase Hi-CDNAse Hi-C complements high resolution Hi-C approach with restriction enzymes.
Hi-CHi-C uses high-throughput sequencing to find the nucleotide sequence of fragments ( all-vs-all).
Hi-C^2Hybrid Capture Hi-C (Hi-C^2) is a technology that combines targeted genomic capture and existing situ Hi-C libraries to observe conformation changes in selected genomic regions.
iMARGIiMARGI (in situ mapping of RNA-Genome Interactome) is a technique that globally maps native RNA-genome interactions from unperturbed cells. It is an improved version of traditional MARGI published in 2017.
In situ Hi-CIn situ Hi-C method is used to evaluate all DNA-DNA proximity ligation in intact nuclei.
Micro-CMicro-C enables mono nucleosome-resolution analysis of chromosome folding fragmentation using micrococcal nuclease (MNase). Micro-C XL is implemented by adding long x-linkers to the fragments.
NLANuclear Ligation Assay (NLA) is an historical method developped in 1993 to determine circularization frequencies of DNA in solution. It inspired 3C method.
PLAC-seqProximity Ligation Assisted ChIP-seq (PLAC-seq), also known as HiChIP, combines in situ proximity ligation with chromatin immunoprecipitation (ChIP) to map chromatin interactions centered on genomic regions bound by the transcription factor or histone with the specific modifications with much reduced cost compared to Hi-C.
Repli-SeqRepli-Seq is a genome-scale approach to map temporally ordered replicating DNA using massively parallel sequencing.
Single-Cell DamIDSingle Cell DamID enables visualization of in vivo with adenine-6-methylation of intact cells by couopling DamID technique and engineered DpnI digestion enzyme.
Sci-Hi-CSingle-cell combinatorial indexed Hi-C (Sci-Hi-C) is a high throughput method that enables mapping chromatin interactomes in large number of single cells.
Single-cell Hi-CSingle Cell Hi-C is an adaptation of Hi-C to single-cell analysis, by including in-nucleus ligation.
Split-pool BarcodingSplit-pool barcoding or Drop-seq is a strategy for profiling thousands of individual cells by separating them into nanoliter-sized aqueous droplets and associating a different barcode with each cell's RNAs and sequencing them all together.
TRIPThis is protocol for analyzing thousands of reporters integrated in parallel (TRIP) at a genome-wide level. TRIP is based on tagging each reporter with a unique barcode, which is used for independent reporter expression analysis and integration site mapping?

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