Emergency Department of The Third Affiliated Hospital of Chongqing Medical University, 1 Huixing Shuanghu Branch Road, Yubei District, Chongqing, 401120, China.
Appl Biochem Biotechnol. 2022 Jul;194(7):3136-3144. doi: 10.1007/s12010-022-03907-7. Epub 2022 Mar 28.
MicroRNAs (miRNAs) play crucial roles in regulating various biological processes and are considered promising biomarkers for clinical diagnosis and therapy of acute pancreatitis. Herein, we present a duplex-specific nuclease (DSN enzyme) and DNAzyme-assisted fluorescent miRNA detections assay that can provide improved detection specificity due to a design of dual-target recognition and a comparable sensitivity. The dual-target recognitions are composed of (i) miRNA unfold hairpin structure toehold to form DNA-RNA duplex, among which the DNA section will be digested by DSN enzyme, releasing miRNA to participant in a next recycle. (ii) After DNAzyme-based nicking site formation in loop section of molecular beacon (MB), miRNA can bind with the loop section of MB and gradually unfold MB probe, generating fluorescence signals. With this general principle, distinct discrimination capability towards even one base pair mismatch of homogenous miRNA is obtained, showing a promising prospect in clinical diagnosis and therapy of acute pancreatitis.
MicroRNAs (miRNAs) 在调节各种生物过程中发挥着关键作用，被认为是急性胰腺炎临床诊断和治疗有前途的生物标志物。在此，我们提出了一种双链特异性核酸酶（DSN 酶）和 DNA 酶辅助的荧光 miRNA 检测分析方法，由于采用了双靶识别设计，其检测特异性得到了提高，并且具有可比的灵敏度。双靶识别由（i）miRNA 展开发夹结构的悬垂部分形成 DNA-RNA 双链，其中 DNA 部分将被 DSN 酶消化，释放 miRNA 以参与下一个循环。（ii）在分子信标（MB）环部分形成基于 DNA 酶的切口后，miRNA 可以与 MB 的环部分结合并逐渐展开 MB 探针，产生荧光信号。基于这一原理，即使是同源 miRNA 的一个碱基错配也能获得明显的区分能力，在急性胰腺炎的临床诊断和治疗中具有广阔的应用前景。
