CTFFIND5 provides improved insight into quality and geometry of cellular cryo-EM samples

Johannes Elferich, Lingli Kong, Ximena Zotttig, Nikolaus Grigorieff

This repository contains the raw text, figures, and scripts used to generate a manuscript describing improvements to the CTFFIND program targeted at cryo-EM imaging of cellular samples.

Get started

Clone this repository

git clone https://github.com/GrigorieffLab/ctffind5_manuscript.git ./

Make sure Git LFS is setup

git lfs install

Setup the conda environment

conda env create -f build/environment.yml

Activate the environment:

source activate ctffind5_manuscript

Build the manuscript:

./build/build.sh

Tools

The following tools are used for this purpose:

• pandoc to convert the markdown files into Word or PDF files
• manubot-cite-pandoc to manage citations
• miniconda to maintain a virtual environment for the required tools
• inkscape to assemble panels into figures
• python and matplotlib to generate plots
• tikz to generate flowcharts
• git-lfs to maintain image files

Files and directories

• manuscript.md the manuscript file.
• output/ contains the rendered manuscript.
• figures/ contains the inkscape .svg files and .png files created from them.
• panels/scripts contains python scripts that create plots.
• panels/images contains the image files resulting from the scripts. IMages could be placed directly in there, but the preference is to have them generated by code.
• scripts/ contains scripts that do data processing, but do not directly result in a panel.

TODO items

• Choose pixel size for fitting better
• Find datasets for tomo vs thickness verification
• Fix OverlayCTF
• Just change FRC threshold?
• Find out why it fails for certain images atm
• Options for
  • McMullan vs CTFFIND style model
  • Reduce weight of node area in fitting

Dataset for tomo thickness verification

Tomos

Pixel Size 4.1746
/data/elferich/CryoTEM/Johannes_20210412/0401_grid10/st011.mrc
/data/elferich/CryoTEM/Johannes_20210412/0401_grid10/st003.mrc
/data/elferich/CryoTEM/Johannes_20210412/0401_grid10/st008.mrc
Pixel Size 2.0873
/data/elferich/CryoTEM/Johanes_20210614/144diff_NP2/s003.mrc
/data/elferich/CryoTEM/Johanes_20210614/144diff_NP1/s001.mrc
/data/elferich/CryoTEM/Johanes_20210614/120diff_Y12/s003.mrc
/data/elferich/CryoTEM/Johanes_20210614/120diff_Y12/s006.mrc

Exposures

Undiff
/data/elferich/CryoTEM/Johannes_20210412/0401_grid10/frames/stack_record_40_00000_-0.0.tif -> 2843A
/data/elferich/CryoTEM/Johannes_20210412/0401_grid10/frames/stack_record_25_00000_-0.0.tif -> 1002A
/data/elferich/CryoTEM/Johannes_20210412/0401_grid10/frames/stack_record_32_00000_-0.0.tif -> 2215A
144
/data/elferich/CryoTEM/Johanes_20210614/144diff_NP2/frames/s_records_20_00000_-20.0.tif -> 2147A
/data/elferich/CryoTEM/Johanes_20210614/144diff_NP1/frames/s_records_29_00000_-20.0.tif -> 1866A
120
/data/elferich/CryoTEM/Johanes_20210614/120diff_Y12/frames/s_record_25_00000_-20.0.tif -> 2383
/data/elferich/CryoTEM/Johanes_20210614/120diff_Y12/frames/s_record_36_00000_-20.0.tif -> 2184

Figures

Figure 1: How does tilt estimation work?

Figure 2: How does thickness estimation work?

Figure 3: CTF correction of low magnification maps

Figure 4: Verification and error estimate of tilt

Figure 5: Verification and error estimate of thickness

Notes

Minimum 600dpi EPS TIFF 8.85cm width