Question: I want to simulate noise and CTF to generate "real" EM-pictures from the projections of my molecule, just as you did in the orginal EMAN-paper for the BTV trimer. When I measure the carbon-film and use ctfit to determine the ctf, are the noise-levels determined simultaneously? How do I apply these parameters to the pictures?

Answer: This feature is not well documented. The best approach is:

1) generate a set of projections 'proj.hed'

2) 'ctfit proj.hed'

3) On the 'Process' menu select 'Simulate' -> 'RT CTF Sim'

• this will open a file dialog, select proj.hed again

4) A window with a picture of your first projection will appear

• as you modify the ctf parameters, the appearance of this image will be updated continuously.

5) Set the desired parameters. A reasonable set for a decent FEG scope is:

• 200 kev, 1 mm Cs Noise : 0, 1, 6, 3 Amp Cont: 10 Envelope: 7.5 Defocus: whatever you want, usu 1-4 microns

6) This will set the basic parameters. The one parameter you cannot define

• a 'typical' value for is 'Amp', since it depends on the amplitude of your projections (usually you'd start out with a normalized 3d model). Basically you adjust 'Amp' to achieve the desired signal to noise ratio visually. Once you've applied the ctf to the entire set of projections, you can read the simulated set back into ctfit again and redetermine the SNR.

7) select 'process' -> 'simulate' -> 'apply ctf 2d'

• This will not overwrite the input file. It will create a new file with the CTF parameters determined

Note: if you want to do this on many files, you can use 'applyctf' once you have a good set of parameters.