Local researchers said they have developed a test method that can simultaneously identify 20 kinds of genes, such as DNA and micro RNA (miRNA), as well as RNA viruses like Covid-19.
The research team led by Dr. Kim Sang-kyung at the Center for Molecular Recognition Research of the Korea Institute of Science and Technology (KIST) said Tuesday it found a new way of detecting genes up to 20 types through multiplex quantitative reverse transcription-polymerase chain reaction (qRT-PCR).
So far, researchers have had technical limitations to analyze five or more genes with qRT-PCR.
In the past, researchers distinguished three or four types of gene signals with different fluorescent colors. However, this method made it difficult to analyze more than five genes at the same time because fluorescent colors could overlap.
To overcome the limitations, KIST’s research team developed a microparticle with a diameter of 500μm that can perform both reverse transcription and PCR. Then, they put discernable patterns in each particle.
The researchers analyzed six genes, including that of influenza, and developed a chip to detect 20 types of genes simultaneously.
The researchers also developed microparticles optimized for analysis of miRNA, a new genetic material that regulates protein output. Chemically, miRNA has the same properties as RNA. Still, its length is very short, making it difficult to design qRT-PCR conventionally, the researchers said.
Dr. Jung Seung-won of KIST immobilized a stem-loop RT primer specifically made for miRNA within the particle for reverse transcription and achieved the PCR reaction in the particle.
This reduced the complex process of miRNA analysis and shortened the testing time to less than one hour.
The process by which RNA is made from DNA is called transcription, and conversely, the process by which complementary DNA is made from RNA is called reverse transcription.
“This study showed that the particle-based diagnostic technology (qPCR) method could be a breakthrough that improves the accuracy and efficiency of testing through multiple sets of genetic information,” Kim said.
The new method can be effectively applied not only to improve the diagnostic accuracy of a single disease with multiple genetic markers but also to accurately identify the source of infection during the outbreaks of various infectious diseases with similar symptoms, he emphasized.
The Ministry of Science and ICT supported the research through the National Research Foundation of Korea’s senior researcher support program and the National Research Council of Science and Technology’s Creative Allied Program (CAP).
The paper, titled, “In-particle stem-loop RT-qPCR for specific and multiplex microRNA profiling,” was published in the latest Biosensors & Bioelectronics issue.