See me, feel me: methods to concurrently visualize and manipulate single DNA molecules and associated proteins.
Direct visualization of DNA and proteins allows researchers to investigate DNA-protein interactions with great detail. Much progress has been made in this area as a result of increasingly sensitive single-molecule fluorescence techniques. At the same time, methods that control the conformation of DNA molecules have been improving constantly. The combination of both techniques has appealed to researchers ever since single-molecule measurements have become possible and indeed first implementations of such combined approaches have proven useful in the study of several DNA-binding proteins in real time. Here, we describe the technical state-of-the-art of various integrated manipulation-and-visualization methods. We first discuss methods that allow only little control over the DNA conformation, such as DNA combing. We then describe DNA flow-stretching approaches that allow more control, and end with the full control on position and extension obtained by manipulating DNA with optical tweezers. The advantages and limitations of the various techniques are discussed, as well as several examples of applications to biophysical or biochemical questions. We conclude with an outlook describing potential future technical developments in combining fluorescence microscopy with DNA micromanipulation technology.
van Mameren
Joost
J
Department of Physics and Astronomy and Laser Centre, VU University, De Boelelaan 1081, 1081 HV, Amsterdam, The Netherlands.
Peterman
Erwin J G
EJ
Wuite
Gijs J L
GJ
D004247:DNA / Q000032:analysis* / Q000737:chemistry; D004268:DNA-Binding Proteins / Q000032:analysis*; D005821:Genetic Techniques; D008856:Microscopy, Fluorescence; D052898:Optical Tweezers
D004268:DNA-Binding Proteins; D004247:DNA