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Urinary exosomal mRNA detection using novel isothermal gene amplification method based on three-way junction

 Jeong Moon  ;  Jaewoo Lim  ;  Seoyoung Lee  ;  Hye Young Son  ;  Hyun Wook Rho  ;  Hongki Kim  ;  Hyunju Kang  ;  Jinyoung Jeong  ;  Eun-Kyung Lim  ;  Juyeon Jung  ;  Yong-Min Huh  ;  Hyun Gyu Park  ;  Taejoon Kang 
 BIOSENSORS & BIOELECTRONICS, Vol.167 : 112474-8, 2020-11 
Journal Title
Issue Date
Cancer diagnosis ; Exosome ; G-quadruplex ; Isothermal gene amplification ; RNA ; Three-way junction
Exosomal messenger RNA (mRNA) has emerged as a valuable biomarker for liquid biopsy-based disease diagnosis and prognosis due to its stability in body fluids and its biological regulatory function. Here, we report a rapid one-step isothermal gene amplification reaction based on three-way junction (3WJ) formation and the successful detection of urinary exosomal mRNA from tumor-bearing mice. The 3WJ structure can be formed by the association of 3WJ probes (3WJ-template and 3WJ-primer) in the presence of target RNA. After 3WJ structure formation, the 3WJ primer is repeatedly extended and cleaved by a combination of DNA polymerase and nicking endonuclease, producing multiple signal primers. Subsequently, the signal primers promote a specially designed network reaction pathway to produce G-quadruplex probes under isothermal conditions. Finally, G-quadruplex structure produces highly enhanced fluorescence signal upon binding to thioflavin T. This method provides a detection limit of 1.23 pM (24.6 amol) with high selectivity for the target RNA. More importantly, this method can be useful for the sensing of various kinds of mRNA, including breast cancer cellular mRNA, breast cancer exosomal mRNA, and even urinary exosomal mRNA from breast cancer mice. We anticipate that the developed RNA detection assay can be used for various biomedical applications, such as disease diagnosis, prognosis, and treatment monitoring.
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1. College of Medicine (의과대학) > BioMedical Science Institute (의생명과학부) > 1. Journal Papers
1. College of Medicine (의과대학) > Dept. of Radiology (영상의학교실) > 1. Journal Papers
Yonsei Authors
Son, Hye Yeong(손혜영) ORCID logo https://orcid.org/0000-0001-5977-6784
Huh, Yong Min(허용민) ORCID logo https://orcid.org/0000-0002-9831-4475
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