Rolling Circle and Loop Mediated Isothermal Amplification Strategy for Ultrasensitive miRNA Detection

Abstract

Rolling circle amplification (RCA) and loop mediated isothermal amplification (LAMP) were combined to establish the rolling circle and loop mediated isothermal amplification (RC-LAMP) method for miRNA detection. With the participation of Bst 2.0 DNA Polymerase, the method enabled RCA and LAMP amplification to occur simultaneously without thermal cycling. The limit of detection of RC-LAMP was 500 amol/L, which is comparable to previously reported amplification strategies. Moreover, its upper limit of quantitation was higher and showed a stronger resistance to matrix interference. Therefore, it is possible to detect low concentrations of miRNA in samples by increasing the total RNA added. Owing to its facile detection mode and simple operation, this method has great potential in clinical sample detection.

Double blocking gap-filling-ligation coupled with cascade isothermal amplification for ultrasensitive quantification of N6-methyladenosine

“…Additionally, to ensure the detection sensitivity and further simplify the requirement for instruments, the gap-filling-ligation reaction is coupled with a cascade isothermal reaction consisting of both rolling circle amplification (RCA) and loop-mediated isothermal amplification (LAMP), which has been well demonstrated in different systems for detecting DNA and RNA with valuable performance. [18][19][20][21] It should be noted that RCA and LAMP are performed in a single tube by using only one kind of polymerase without any other steps.…”