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Comparison of Quantitative PCR and Droplet Digital PCR for Effective Hepatitis B Covalently Closed Circular DNA Measurement |
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| รหัสดีโอไอ | |
| Title | Comparison of Quantitative PCR and Droplet Digital PCR for Effective Hepatitis B Covalently Closed Circular DNA Measurement |
| Creator | Suparerk Borwornpinyo |
| Contributor | Pattida Kongsomboonchoke |
| Publisher | The Thai Society for Biotechnology |
| Publication Year | 2562 |
| Keyword | Covalently Closed Circular DNA, Quantitative PCR, Droplet Digital PCR |
| Abstract | Chronic hepatitis B virus (HBV) infection is caused by the persistence of viral covalently closed circular DNA (cccDNA) in infected hepatocytes. Targeting cccDNA is a novel approach for chronic HBV infection treatments. Since the number of cccDNA presented in a hepatocyte is very low, the precise and sensitive technique has to be developed in order to detect and quantify cccDNA. Even though quantitative polymerase chain reaction (PCR) was used for almost 3 decades by fluorescent dyes or probes to detect the amplification of DNA target, this technique only provides a relative quantification that is dependent with calibration curve. This limitation was resolved by developing of droplet digital PCR (ddPCR). ddPCR offers an absolute quantification that calculated based on Poisson algorithm. Furthermore, it is advantageous by high sensitivity and reproducibility to detect low amplicon template with extreme precision. This study presents the successful validation of ddPCR for applying to estimate cccDNA copy number. By using known concentration of plasmid, ddPCR showed the lowest detection limit around 0.1 copies per 1X reaction volume. The accuracy to quantify cccDNA target was compared between theoretical and practical value and determined the correlation by R-squared close to 1. From our results, we exhibited that ddPCR should be a platform for quantifying cccDNA and applied in subsequent researches in order to evaluate the precise cccDNA number after cccDNA-targeting treatments |
| Language | EN |
| URL Website | http://tsb2019.com/ |
| Website title | The 31st Annual Meeting of the Thai Society for Biotechnology and International Conference (TSB 2019) |
