Establishment of a dual-fluorescent MSI reporter system

• Main Authors: Chien-Yuan Chiu, 邱建元
• Other Authors: Christina Ling Chang
• Format: Others
• Language: en_US
• Published: 2009
• Online Access: http://ndltd.ncl.edu.tw/handle/67904038637685998940

碩士 === 國立成功大學 === 口腔醫學研究所 === 97 === Microsatellites are simple repeats of 1-6 nucleotides, which are widely dispersed in the human genome. Deficiency in DNA mismatch repair (MMR) system results in unrepaired replicative errors preferably in microsatellite sequences, leading to microsatellite instability (MSI). The MSI phenotype has been detected in many other cancers, such as colon and oral cancers. Clinically, cancer patients with MSI develop resistance to chemotherapy. Therefore, it is important to screen and develop anti-MSI compounds in order to reduce cancer incidence and mortality. I have constructed expression plasmids containing (CA)n microsatellite sequences to report the MSI phenotypes in a dual-fluorescence fashion. Since oxidative stress inactivates the MMR function, I have examined the sensitivity of different microsatellites in reporting H2O2–induced MSI frequency in transient and stable transfectants derived from HCT116 and HCT116+chr3 human colorectal cancer cell lines. With flow cytometry, I found that H2O2 increased the MSI frequency in a dose-dependent manner in stable transfectants that harbor (CA)5 and (CA)13 microsatellites, but not in transient transfectants. Furthermore, H2O2 also increased the MSI frequency in a time-dependent manner in the stable transfectants. Since little is known about the correlation between MSI and drug resistance in oral cancer, I tested and found that OSCC25 cells displayed higher drug resistance than CAL27 cells. This is consisted with our previous finding that the MSI frequency is higher in OSCC25 than CAL27 cells. To control and prevent cancer, the dual fluorescent MSI reporters that I have developed will be useful in screening anti-MSI compounds.