Table 2 Common tools to screen and study enhancers

Chromatin features and histone modifications

ChIP-seq,

CUT&RUN,

CUT&Tag

Can be used to screen putative enhancers in the genome-wide and identify active enhancers. Study chromatin regulation and identify the genome-wide binding sites of specific TFs. CUT&RUN and CUT&Tag, as upgrades of ChIP, requires few cells, CUT&Tag is for single cell

[72, 73, 77, 78]

Chromatin accessibility

ATAC-seq,

DNase-seq,

MNase-seq,

FAIRE-seq

Genome-wide annotation of regulatory regions based on chromatin accessibility for screening of potential enhancers and transcription factors. MNase-seq is a method that reflects chromatin accessibility indirectly, and the other three methods reflect chromatin accessibility directly

[83]

Transcriptional assays

RNA-seq,

CAGE,

GRO-seq

Annotation of active enhancers by eRNA expression levels generated by enhancer transcription. CAGE and GRO-seq methods improved the sequencing sensitivity

[85, 87, 89]

Massively parallel reporter assays

MPRA,

STARR-seq

Can be used to readout enhancer activity on a large scale and understand the functional sequence of enhancers in depth

[92, 95]

Chromosome conformation assays

Hi-C,

ChIA-PET,

HiChIP

Can be widely used to explore enhancer-promoter connections from the 3D architecture of chromosomes and identify target genes of enhancers

[98,99,100]

CRISPR system

CRISPR screens, CRISPR/Cas9, CRISPRa,

CRISPRi

A high-throughput tool for reading out functional enhancers using CRISPR screens.

Functional annotation of potential enhancers and analysis of regulatory mechanisms can be performed using CRISPR/Cas9 and its derivative technologies

[110, 125]

Oligonucleotide method

LNAs

Targeting eRNA to inhibit enhancer activity

[127]