Process movies

As of version 1.2, RELION incorporates functionality to process movies from fast digital cameras, such as direct-electron detection devices (DDDs). The following has been tested on several data sets from our FEI Falcon camera. Other detectors have not been tested yet. Please share your experiences with us if you have.

Get organised

Organize all your movies and the average micrographs (i.e. single-image files that are the average of all frames of your individual movies) inside one or more directories that are inside your RELION project directory (from where you would launch the GUI). We like to call these directories "Micrographs/" if all micrographs are in one directory, or "Micrographs_15jan13/" and "Micrographs_23jan13/" if they are in different directories (e.g. because they were collected on different dates. But, you can organise yourself as you like.

Name the average micrographs and movies in the following way:

001.mrc
001_movie.mrc
24.01.13.15.04.mrc
24.01.13.15.04_movie.mrc
myverybest.mrc
myverybest_movie.mrc
...

Note that all average micrographs and movies are in MRC format with an .mrc extension, and all movies are MRC stacks. (You can also give movies a .mrcs extension, as in RELION's convention). Movies need to have the same rootname (e.g. 001, or 24.01.13.15.04, or whatever you like) as their corresponding average micrographs, plus a movie-identifier (in this case "movie") that follows the rootname after an additional "_" character.

What to first

First, use the average micrographs to extract "normal" particle stacks, and follow the Recommended_procedures for 3D refinement.

Preprocessing movies

Then, extract (box out) the individual movie-frames of all particles from the original micrograph movies. To do this, go back to the Preprocessing run-type in the RELION GUI, and:

  * Keep everything on the I/O tab as it was when you extracted the "normal" particles   
  * Set "Run Ctffind3?" to No
  * Set "Extract particles from micrographs?" to Yes
  * Set "Use movies instead of micrographs?" to Yes
  * Set "Rootname of movie files" to the movie-identifier mentioned above (in the example above "movie")
  * Set "Number of frames to average?" to any desired number from 1 to the half the number of frames in your movies
  * Keep everything on the operate tab as it was when you extracted the "normal" particles   
  * Let it run... !

(By this time you will probably already be aware of disc quota, full discs, etc....) For each movie, a separate stack with all movie frames of all particles in that micrograph will be stored along the extracted "normal" particle stacks in the Particles directory. All metadata regarding these particle-movies will be stored in a large STAR file in your project directory that is called particles_movie.star (where "particles" is the Particle rootname given on the I/O tab; and "movie" is your movie-identifier).