As part of the Imaging ONEWORLD series, the focus of these lectures is on microscopy and image analysis methods and how to apply these to your research. Almost all aspects of imaging such as sample preparation, labelling strategies, experimental workflows, ‘how-to’ image and analyse, as well as facilitating collaborations and inspiring new scientific ideas will be covered. Speakers will be available for questions and answers. The organisers, core facility staff from the University of Cambridge, Gurdon Institute, MRC-LMB and the ICR/Royal Marsden Trust are also able to continue the discussion and provide advice on your imaging projects.

Scientific Organisers

Exploring gene transcription mechanisms using single-molecule fluorescence imaging in vitro and in vivo 

Single-molecule studies offer unprecedented and direct access to biologically important heterogeneity and dynamics, and provide dynamic views of biological machines at work; this holds especially true for reactions inside the complex biological milieu of living cells. During the past few years, we have developed and used a wide variety of in vitro and in vivo single-molecule fluorescence methods (single-molecule FRET, super-resolution imaging, single-particle tracking) to answer long-standing questions in gene transcription and DNA repair. Here, I will discuss examples of applications of such methods to unravel the mechanisms of bacterial gene transcription. I will first discuss our studies using single-molecule FRET and unwinding-induced fluorescence enhancement studies to elucidate the intricate sequence of conformational changes that allow RNA polymerase to open the double-stranded promoter DNA for transcription initiation. I will then discuss examples of our in vivo single-molecule work inside living bacteria, focusing on the exploration of the interplay between chromosome organization, gene expression, and bacterial growth.