Ligation-free isothermal nucleic acid amplification

Advances in Simple, Rapid, and Contamination-Free Instantaneous Nucleic Acid Devices for Pathogen Detection

“…Nucleic acid testing consists of three main steps: nucleic acid extraction [34][35][36][37][38][39][40][41][42][43][44], nucleic acid amplification [45][46][47][48][49][50][51][52][53][54][55][56][57], and detection methods [58][59][60][61][62]. The general procedure for nucleic acid extraction involves collecting the sample to be tested (such as oropharyngeal swabs, fingertip blood, or saliva) and then obtaining the nucleic acid through a process of lysis, separation, precipitation, and purification [63][64][65].…”

CRISPR/Cas12a-Powered Amplification-Free RNA Diagnostics by Integrating T7 Exonuclease-Assisted Target Recycling and Split G-Quadruplex Catalytic Signal Output

“…RNAs, as genetic massagers in biological cells, play an irreplaceable role in gene coding and regulation across almost all life processes. , Its great value in disease diagnosis and prognosis assessment has also attracted considerable interest in recent years . For instance, bacterial 16S rRNA (rRNA), which is highly conserved between different bacterial species and relatively abundant in viable cells, serves as an ideal target in species-specific pathogen identification. − Noncoding RNAs, including MicroRNAs and long noncoding RNA (lncRNA), are another important kind of RNA that participate in life activities as “organizers” and regulatory molecules, and their aberrant expression has become an important diagnostic and prognostic biomarker for malignant tumors, cardiovascular diseases, and nervous system diseases. , Although traditional methods, such as reverse transcription-quantitative polymerase chain reaction (RT-qPCR), Northern blots, and microarray analysis, have been well established for RNA detection, the involvement of tedious process, dedicated equipment, and trained personnel largely limited their applications to centralized laboratories. , Isothermal amplification techniques, including nucleic acid sequence-based amplification (NESBA), recombinase polymerase amplification (RPA), and loop-mediated isothermal amplification (LAMP), avoid complex thermal cycling and shorten the detection time. − However, their applications are still restricted by the sophisticated primer design and the risk of aerosol contamination. Therefore, a rapid, simple, and sensitive method for RNA detection remains an urgent need.…”