From 2010.igem.org
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ApoControl
The control of gene expression in eukaryotic systems has been challenging due to the complexity of the structure and the physiology of eukaryotic cells. The project ApoControl is focused on developing a system to improve the efficiency and specificity of gene expression in eukaryotic cell-lines. Here proapoptotic gens with inducible promoters, instead of antibiotic resistance, are used as selection marker to induce clean cell-death at appropriate stimuli. The project is divided into three parts, each representing different approaches to achieve this goal.
Cut'N'survive System
This system utilizes the property of N-Degron to degrade the proapoptotic protein when target gene is successfully transfected into the cell. The recognition and activation of N-Degron is carried out by TEV-protease. Thus this system enables selection of cells expressing the target gene while the cells not transfected undergo induced apoptosis.
Jump-or-Die System
This system makes use of the PhiC31o integrase and corresponding recombining sequences to integrate a target gene into one specific site in the chromosome of the cell line. Cells with integrated target gene will not express the proapoptotic gene due to the stop codon after the target gene while other cells undergo induced apoptosis.
The ProSearch System
Inducible systems in eukaryotic cells can be highly valuable both in science and in clinic if there are a wide variety of environmental conditions that can promote target gene expression. The aim of this system is to search for new enhancer-promoter combinations that would efficiently trigger gene expression in response to different stimuli. This part of the project is carried out through bioinformatic modeling process. The end-product will be a databank of related genes and translation factors at the input of wished environmental conditions (hypoxia, hyperthermia etc) and the used cell-line. This system will be useful for the construction of new inducible systems trying to utilize apoptosis or other gene expression in human cell-lines.
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