Lab Course: Mammalian Gene Circuits

Abstract

The students are trained in basic techniques in construction and characterization of synthetic gene circuits in mammalian cells. Experimental circuits are built with both the input and the output conjugated to fluorescent reporters, allowing characterization at the single cell level.

Objective

The objective of the course is to construct a genetic sensor for a molecular regulatory input such as microRNA or a transcription factor and characterize the input/output relationship of this sensor with the help of fluorescent reporters, fluorescent microscopy and fluorescent-activated cell sorting. The emphasis is on single-cell characterization.

Content

The course will take place over 4 weeks, with 2 days per week spent on lab work. The 4 weeks will be dedicated to the following activities Week 1: Introduction to the course; supervised construct design and detailed planning. Cloning of the constructs: part 1. Week 2: Cloning of the constructs, purification and characterization of DNA constructs Week 3: Cell culture transfection, microscopy and flow cytometry characterization Week 4: Data analysis and preparation of the final report; possibility to repeat failed experiments.

Resources