Table 1.

List of techniques to investigate RNA matchmaking interactions.

Direct Interactions with DNA
Method	Concept	Reference
ChOP	Modified ChIP assay that uses biotinylated antisense oligonucleotide to affinity purify the target RNA and associated chromatin (PCR is used to identify binding at specific regions of the genome)	Mariner PD et al., 2008
ChIRP	Uses multiple tiling affinity-tagged antisense oligos and glutaraldehyde (DSG) crosslinking to pulldown a targeted RNA with its associated DNA (chromatin is then subjected to deep sequencing)	Chu C et al., 2011
CHART	Similar to ChIRP, uses formaldehyde instead of DSG and only uses RNase H (as opposed to the cocktail of RNase A and H used in ChIRP) to specifically elute RNA interacting with chromatin	Simon, MD et al., 2011
RAP	Similar to ChIRP and CHART, uses formaldehyde and omits the RNase treatment in favor of DNase to digest the genomic DNA to ∼150bp sized fragments which provides high-res mapping of binding sites	Engreitz JM et al., 2013
CHIRT-seq	Similar to ChIRP and CHART, but only uses a single oligo probe to allow for profiling of more repetitive RNAs	Chu HP et al., 2017
MARGI-seq	Genome-wide approach that captures all RNA:DNA interactions by coupling proximity ligation with a unique linker sequence that allows the RNA and DNA components to be differentiated	Sridhar B et al., 2017
GRID-seq	Similar to MARGI-seq, but uses 2-step crosslinking to capture longer-range RNA:DNA interactions and a restriction enzyme step to create identical size-selected fragments	Li X et al., 2017
ChAR-seq	Similar to MARGI-seq and ChAR-seq, uses long single-end reads to accurately capture the polarity of the bridge linker and performs the proximity ligation on intact nuclei to reduce spurious events	Bell JC et al., 2018
Hi-ChIRP	Combines ChIRP-seq with Hi-C chromatin conformation capture to characterize specific RNAs involved in mediating inter-chromosomal interactions	Mumbach MR et al., 2019
RADICL-seq	Similar to GRID-seq, but uses fewer cells, omits the 2-step crosslinking, utilizes DNase 1 instead of restriction enzyme to partially digest chromatin, and generates longer fragments to increase mapping efficiency	Bonetti A et al., 2020
RD-SPRITE	Modification of the split-and-pool based SPRITE technique that couples crosslinking with a unique barcoding strategy to examine the three-dimensional organization of interacting RNA and DNA within the nucleus	Quinodoz S et al., 2020
RNA-RNA Interactions on Chromatin
Method	Concept	Reference
RPL	Uses proximity ligation followed by deep sequencing to profile global RNA:RNA interactions, though without crosslinking is largely limited to profiling intra-molecular RNA interactions	Ramani V et al., 2015
RAP-RNA	Modified RAP method, uses affinity-tagged antisense oligos and UV crosslinking of the psoralen-based crosslinker aminomethyltrioxalen (AMT) to directly capture interactions mediated by a target RNA	Engreitz JM et al., 2014
LIGR-seq	Uses in vivo crosslinking with AMT, proximity ligation, and 2D purification of RNA duplexes to generate genome-wide maps of RNA:RNA interactions	Sharma E et al., 2016
PARIS	AMT crosslinking coupled with proximity ligation, uses partial RNase digestion to ensure identified crosslinks are limited to direct base-paired RNAs and 2D electrophoresis to purify only crosslinked fragments	Lu Z et al., 2016
SPLASH	Use biotinylated psoralen to crosslink RNA before undergoing proximity ligation and deep sequencing	Aw JG et al., 2016
COMRADES	Uses in vivo crosslinking of an azide-modified psoralen derivative and biotinylated DNA oligos to selectively capture interactions between a target RNA and proximal RNA molecules	Ziv O et al., 2018
Proximity RNA-seq	Uses a water-in-oil emulsion barcoding technique followed by high-throughput sequencing to profile the subcellular localization of RNA interactions within the nucleus	Morf J et al., 2019
RNA Interactions Mediated by RNA Binding Proteins
Method	Concept	Reference
CLASH	Uses UV crosslinking, ligation, and immunoprecipitation to enrich for interacting RNA molecules mediated by a specific RNA binding protein (RBP) and sequencing the resulting hybrids	Helwak A et al., 2013
hiCLIP	Similar to CLASH, identifies RNA duplexes by ligating two interacting RNAs but incorporates an adaptor between the two strands to increase ligation efficiency and the ability to identify the respective molecules	Sugimoto Y et al., 2015
MARIO	Global profiling of RNA:RNA interactions via UV crosslinking of RNAs to biotinylated RBPs followed by proximity ligation to link RNA molecules that are associated with the same protein	Nguyen TC et al., 2016
RIC-seq	Uses a biotinylated cytidine phosphate (pCp-biotin) linker to label RNAs and then captures RNA:RNA interactions via proximity ligation at single-nucleotide resolution)	Cai Z et al., 2020