title: Ultrasensitive detection of microRNAs based on hairpin fluorescence probe assisted isothermal amplification.
abstract: A hairpin fluorescence probe assisted isothermal amplification strategy was used for microRNAs (miRNAs) detection. The fluorescence hairpin probe was rationally designed by software NUPACK to reduce background signal. This isothermal amplification method consisted of two circuits. The amplification strategy not only could detect miRNA, but also amplified and reversely transcribed miRNA into DNA to enhance the stability of the target. The approach was ultrasensitive and as low as 8.5×10(-15)mol/L miR-Let-7a, corresponding to 8.5×10(-20)mol miR-Let-7a in 10µL, was able to be detected within 20min at 37°C. Moreover, successful detection of miR-Let-7a in a total RNA sample was also achieved. Thus, the rapid, simple, isothermal, and highly sensitive approach should be a promising tool for on-the-spot detection.
authors.affiliation: State Key Laboratory Base of Eco-chemical Engineering, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao 266042, PR China.
authors.forename: Cuiping
authors.initials: C
authors.lastname: Ma
authors.affiliation: State Key Laboratory Base of Eco-chemical Engineering, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao 266042, PR China.
authors.forename: Sen
authors.initials: S
authors.lastname: Liu
authors.affiliation: State Key Laboratory Base of Eco-chemical Engineering, College of Chemistry and Molecular Engineering, Qingdao University of Science and Technology, Qingdao 266042, PR China. Electronic address: sc169@163.com.
authors.forename: Chao
authors.initials: C
authors.lastname: Shi
mesh_terms: D001483:Base Sequence; D015374:Biosensing Techniques / Q000295:instrumentation*; D004867:Equipment Design; D019544:Equipment Failure Analysis; D005456:Fluorescent Dyes / Q000737:chemistry*; D035683:MicroRNAs / Q000032:analysis* / Q000737:chemistry / Q000235:genetics; D008832:Microchemistry / Q000295:instrumentation*; D008969:Molecular Sequence Data; D021141:Nucleic Acid Amplification Techniques / Q000295:instrumentation*; D013050:Spectrometry, Fluorescence / Q000295:instrumentation*; D013696:Temperature
chemical_list: D005456:Fluorescent Dyes; D035683:MicroRNAs
keywords: Cascade amplification; Hairpin probe; Isothermal amplification; MicroRNAs; Real-time fluorescence