Related Experiment Videos
Assessing optimal promoter activity for constructs in gastrointestinal gene therapy
Rabea Mathlouthi1, Susanne Aberle, Nicola Schug
1Division of Molecular Genetics, University of Tübingen, Wilhelmstr 27, D-72074 Tübingen, Germany.
Background:
During recent decades, studies of various cancer-related genes has led to a growing understanding of molecular mechanisms of gastrointestinal cancer resulting in a genetic progression model. Nevertheless, with a few exceptions, our knowledge of participating genes has not been exploited for gene therapeutic approaches. Therefore, we monitored the promoter activity of genes shown to be significantly expressed in gastrointestinal tumors to select optimally active promoters for recombinant DNA constructs. Such molecules will contain a suicide gene under a suitable cell type-specific regulatory element.
Materials And Methods:
Using promoter-reporter gene (luciferase) constructs we compared the activities of KRT19, TFF1, SEL1L, MUC4, MUC1, CEL and hTERT by transfecting them into the gastrointestinal cell lines MKN45 and DAN-G for transient expression. Furthermore we tested the endogenous expression of these genes by RT-PCR.
Results:
From a selection of 9 promoter constructs SEL1L, KRT19 and MUC1 displayed the highest activity levels while others were moderately expressed.
Conclusion:
These three appear the most capable ones to drive suicide genes, like thymidine kinase or cytosine deaminase, a study presently initiated.