Lab of Jeff Gelles
Department of Biochemistry at Brandeis University

Home

Research

Techniques

Movies

People

Lab Photos

Collaborators


Software

Plasmids


Brandeis Lab Page

Movies

Transcription against an Applied Force
[MPEG format (0.88 Mb)] [MPEG format (with superimposed graphics, 1.47 Mb)]
[SGI Movie format (1.04 Mb)] [SGI Movie format (with superimposed graphics, 1.48 Mb)]
In case of difficulty: How to view an MPEG movie using your web browser

This movie shows an experiment from:
"Transcription against an Applied Force"
Hong Yin, Michelle D. Wang, Karel Svoboda, Robert Landick, Steven M. Block, and Jeff Gelles
(1995) Science 270, pp. 1653-1657. [abstract]


The movie is a video recording, made through the microscope, of the experiment that produced the data in Fig. 2A of the paper. The movie shows a DNA-tethered bead exhibiting tethered Brownian motion, being captured by the laser trap, and being moved by the bound RNA polymerase molecule. Since this experiment was performed at relatively low laser power (25 mW), the polymerase does not stall.

Sequence of events visible in the movie:

  1. Brownian motion of a DNA-tethered bead. The differential interference contrast optics used in the microscope image the single spherical bead as apposed black and white ovoids on a gray background. The DNA tether (invisible) confines the random Brownian motion of the bead to a region near the center of the frame.


  2.  
     

  3. Capture and movement of the bead by the laser trap. When the trap is activated by opening the laser shutter, an image of the laser beam appears. The beam image, which consists of a central bright spot and surrounding interference fringes, is produced by a weak reflection of the beam at the cover slip surface. The bead image is displaced somewhat downward and to the right of the true trap position.

  4.  

    When the bead is captured by the trap, the visible Brownian motion stops. The trap center is then moved toward the upper right corner of the frame, dragging the bead with it and placing the DNA tether under light tension.
     
     

  5. Translocation of the bead by the DNA-bound RNA polymerase molecule. As transcriptional elongation occurs, the force exerted on the DNA by the RNA polymerase molecule moves the bead away from the trap center and towards the point at which the polymerase molecule is attached to the coverslip. This movement is visible in the movie as a slow displacement of the bead towards the lower left corner of the frame. Also visible are the periodic movements of the laser beam in the same direction; these movements serve to keep the bead positioned within the central, calibrated portion of the laser trap.
Events 1 and 2 are shown in real time. The portion of the movie showing event 3 is speeded up twenty-fold in order to make the RNA polymerase-driven bead movement more visible

 The width of the frame is ~3200 nm.

 A second version of the movie is identical to the first except that it has superimposed graphics. In these graphics, the number in the upper left corner of the frame indicates the time acceleration. During event 3, a new white line that points to the trap center appears each time the trap is repositioned.


Many thanks to Daniel Peisach for help constructing the movie.


Background photo credit: Diana Hunt. Last modified 1/28/2010 by Margaret Fairman Williams