Table 2 G4 detection methods and their characteristics

Prediction

Regular expression matching

The potential G4 sequence can be predicted according to the nucleic acid sequences

Dynamic eG4s cannot be predicted by existing prediction tools

Quadparser, Quadruplexes

[4, 5]

Scoring

QGRS mapper, pqsfinder

[6, 7]

Sliding window

G4P calculator, G4Hunter

[8, 9]

Machine learning

Quadron

[10]

Detection and characterization

Biochemical method

The location of G4s in the background of long-chain nucleic acid sequence can be determined

It is impossible to observe G4 dynamics, and most techniques can only detect G4s in vitro

Polymerase stop, DMS and piperidine cleavage

[15, 16]

Biophysical method

It can characterize the structure of G4 and present G4 dynamics in vitro

Only G4s in short-chain nucleic acid sequences can be detected, and G4s can only be detected in vitro

CD, NMR

[11, 12]

Cell imaging

The dynamics of eG4s and the intracellular factors affecting eG4 dynamics can be studied in vivo

Not all individual eG4s in the cells can be detected, the resolution needs to be improved, and the detected eG4s lack sequence information

G4 antibody immunofluorescence, G4 small molecular fluorescent probes

[23,24,25,26,27,28,29,30,31]

Mapping based on NGS sequencing

Polymerase stalling approach

The potential G4 in the genome or transcriptome can be comprehensively detected and the G4 map of the whole genome or transcriptome of the species can be obtained

The operation object is the purified genomic DNA or total mRNA, the environment is artificially controlled, and thus the detection results do not represent the actual eG4s in vivo. It is impossible to study the effect of cellular factors on the G4 dynamics

G4-seq

[17]

Antibody based pull-down approach

The operation object is fixed chromatin (G4 ChIP-seq) or native cells (G4 CUT&Tag). Genome-wide pre-existing eG4s can be detected in vivo. The effect of cellular factors on eG4 dynamics can be studied

The specificity of antibody binding to eG4s still needs to be improved

G4 ChIP-seq, G4 CUT&Tag

[32, 34]