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Urinary exosomal mRNA detection using novel isothermal gene amplification method based on three-way junction.
Jeong Moon1, Jaewoo Lim2, Seoyoung Lee3
1Bionanotechnology Research Center, KRIBB, Daejeon, 34141, Republic of Korea; Department of Chemical and Biomolecular Engineering (BK 21+ Program), KAIST, Daejeon, 34141, Republic of Korea.
Biosensors & Bioelectronics
|August 18, 2020
Summary
This study introduces a rapid, one-step method for detecting exosomal messenger RNA (mRNA) using isothermal gene amplification. The assay shows high sensitivity and selectivity, enabling potential applications in disease diagnosis and monitoring.
Area of Science:
- Biochemistry
- Molecular Biology
- Biotechnology
Background:
- Exosomal messenger RNA (mRNA) is a promising biomarker for liquid biopsy due to its stability and regulatory roles.
- Accurate and sensitive detection of exosomal mRNA is crucial for disease diagnosis and prognosis.
Purpose of the Study:
- To develop a rapid, one-step isothermal method for detecting exosomal mRNA.
- To establish a sensitive and selective assay for urinary exosomal mRNA from tumor-bearing mice.
Main Methods:
- Utilized a three-way junction (3WJ) formation for isothermal gene amplification.
- Employed DNA polymerase and nicking endonuclease for signal primer amplification.
- Integrated G-quadruplex formation and thioflavin T for enhanced fluorescence detection.
Main Results:
- Achieved a detection limit of 1.23 pM (24.6 amol) with high selectivity for target RNA.
- Successfully detected urinary exosomal mRNA from tumor-bearing mice.
- Demonstrated versatility for sensing various mRNA types, including breast cancer-associated mRNA.
Conclusions:
- The developed assay offers a rapid and sensitive platform for exosomal mRNA detection.
- This method holds potential for diverse biomedical applications, including disease diagnosis, prognosis, and treatment monitoring.
