Single-particle tutorial (EMPIAR-10025)

This tutorial shows how to convert raw movies from EMPIAR-10025 (T20S proteasome) into a ~3A resolution structure.

Total running time required to complete this tutorial: 45 min.

We first use the command line to download and decompress a tbz file containing a subset of 20 movies, the gain reference, and an initial model:

# cd to a location in the shared file system and run:

tar xvfz nextpyp_spr_tutorial.tbz

Open your browser and navigate to the url of your nextPYP instance (e.g.,

Step 1: Create a new project

Step 2: Import raw movies


Click inside the Single Particle (from Raw Data) block to see a larger version of the image

Step 3: Pre-processing


While on the Micrographs tab, use the navigation bar at the top of the page to look at the results for other micrographs

Step 4: Reference-based refinement

Step 5: Filter bad particles

Step 6 Permanently remove bad particles

Step 7: Particle refinement


Use the navigation bar at the top left of the page to look at the results for different iterations

Step 8: Create shape mask

Step 9: Local refinement

Step 10: Particle-based CTF refinement

Step 11: Movie frame refinement

Step 12: Refinement after movie frame refinement

Step 13: Map sharpening


Running times were measured running micrographs in parallel on nodes with 124 vCPUs, 720GB RAM, and 3TB of local SSDs