Ultrasensitive detection of telomerase activity in a single cell using stem-loop primer-mediated exponential amplification (SPEA) with near zero nonspecific signal

Abstract

A SPEA strategy is developed for the detection of telomerase activity in a single cell with a near zero nonspecific signal.

Advances in the detection of telomerase activity using isothermal amplification

“…After isothermal amplification, target induced fluorescence emission change can be easily detected even by naked eye 64 . Recently, stem-loop primer-mediated exponential isothermal amplification strategy reported by Li et al was used for accurately and specifically detecting telomerase activity 65 . By strategically designing two stem-loop structures, the assay can nonspecifically detect telomerase extracted from single cell.…”

One-pot detection of telomerase activity with high sensitivity and specificity via RNA FRET probes and RNase H-assisted signal cycling amplification

“…Comparison between the proposed one-pot assay and other reported methods 13,15,17,21,22,24,25,29,30 for the detection of telomerase activity is shown in Table S1 in ESI. † The table reveals that present assay is among the sensitive analysis schemes of telomerase.…”

“…In 2016, we developed a stem-loop primer-mediated exponential amplication (SPEA) strategy for accurately and specically detecting the telomerase-elongated telomere repeats with near zero nonspecic signal. 29 The SPEA-based assay can detect telomerase activity in a single cell with one-type of DNA polymerase under isothermal conditions. Although the sensitivity has been pushed to extremely low detection limits, this method requires DNA amplication with a complex design, using two probes and two primers.…”

Programmable Biosensors Based on RNA-Guided CRISPR/Cas Endonuclease

“…Indeed, Sp Cas9 provides the cleavage function as a novel on-off tool. First, nicking triggered exponential amplification reaction (NTEXPAR) exhibits a versatile and highly efficient pre-screening of sgRNAs to analyze Sp Cas9 cleaved dsDNA in genome editing [ 39 , 40 ]. Subsequently, improvement of NTEXPAR combined with Cas9, with cleavage started by PAMmer (the design of a single antisense PAM oligonucleotide) without the limits of PAM sequence, was successfully developed to detect DNA methylation and L. monocytogenes total RNA; this strategy obtained a detection limit of 0.82 amol and specificity in the recognition of single-base mismatch, as well as providing a new paradigm for molecular diagnostic.…”