problem with ctffind4 on mrc movie stack

Submitted by Antje on Fri, 2015-07-24 07:30

Forums

ctffind & ctftilt

Hi Alexis,

I tried to call ctffind4 from the
command line on an mrc movie stacks with approx. 60 frames and faced the following
problem.

When I averaged over one frame, it just calculated the ctf for the first frame, when I
averaged over more frames it always gave me an output like this:

----------------------------------
Working on micrograph 1 of 1

100% [==============================] done!

**warning(Image::ComputeRotationalAverageOfPowerSpectrum): unable to
rescale 1D average of experimental power

Total execution time : 35 seconds

2015-07-22 15:40:24: Fatal error (ImageFile_Init): File format not
supported yet
----------------------------------

Are there special requirements for the .mrc stack? Or do you have an idea, what might be
going wrong?

Many thanks in advance for help and ideas,
Antje

Alexis Fri, 2015-07-24 07:42

Hi Antje,
Sorry you're having this problem.
To help understand what's going wrong, could you please give me:
- the version of ctffind you're running
- the exact input into CTFFind (what you type at the command-line and your answers to the user-interaction questions) and/or the script you use to run it
- the output from the "header" command (from IMOD) run on your input stack of frames (or the equivalent command from MRC, Relion or whichever package you like).. Something that prints out MRC format information, file dimensions etc

Also, does the output filename (for the diagnostic image) already exist? If so, perhaps try to delete it first.

Thanks
Alexis

Antje Fri, 2015-07-24 11:53

Hi Alexis,

thank you for your quick reply
- the version we are using is : ctffind-4.0.16
- the output files did not exsist beforehand
- to try it out we used the interactive command line version, here is an eample of a run, which didn't work (i tried different numbers for Number of frames to average together but the error was always the same)
----------------------------------------------------
----------------------------------------------------
>> /fs/path2ctffind4.0.16/ctffind

** Welcome to CTFFind **
Version - 4.0.16

Input Mode: Interactive
Date & Time: 2015-07-22 13:29:28

Copyright 2015 Howard Hughes Medical Institute. All rights reserved.
Use is subject to Janelia Farm Research Campus Software Copyright 1.1
license terms ( http://license.janelia.org/license/jfrc_copyright_1_1.html )

Input image file name [stack4ctffind4.mrc] : stack4ctffind4.mrc
Input is a movie (stack of frames) [yes] : yes
Number of frames to average together [1] : 62
Output diagnostic filename [ctff4_out_avg1.mrc] : ctff4_out_avg62
Pixel size [1.4] :
Acceleration voltage [300.0] :
Spherical aberration [2.7] :
Amplitude contrast [0.07] :
Size of power spectrum to compute [256] :
Minimum resolution [30.0] :
Maximum resolution [5.0] :
Minimum defocus [5000.0] :
Maximum defocus [50000.0] :
Defocus search step [500.0] :
Expected (tolerated) astigmatism [100.0] :
Find additional phase shift? [no] :

Summary information for file stack4ctffind4.mrc
Number of columns, rows, sections: 4096 4096 62
MRC data mode: 2
Bit depth: 32
Pixel size: 1.000 1.000 1.000

Working on micrograph 1 of 1

100% [==============================] done!

Total execution time : 8 seconds

2015-07-22 13:30:25: Fatal error (ImageFile_Init): File format not supported yet
----------------------------------------------------
----------------------------------------------------
- this is the header information given by the IMOD header function:
>> header stack4ctffind4.mrc

RO image file on unit 1 : stack4ctffind4.mrc Size= 4063233 K

Number of columns, rows, sections ..... 4096 4096 62
Map mode .............................. 2 (32-bit real)
Start cols, rows, sects, grid x,y,z ... 0 0 0 4096 4096 62
Pixel spacing (Angstroms).............. 1.000 1.000 1.000
Cell angles ........................... 0.000 0.000 0.000
Fast, medium, slow axes ............... X Y Z
Origin on x,y,z ....................... 0.000 0.000 0.000
Minimum density ....................... 0.0000
Maximum density ....................... 4783.2
Mean density .......................... 1070.3
tilt angles (original,current) ........ 0.0 0.0 0.0 0.0 0.0 0.0
Space group,# extra bytes,idtype,lens . 0 0 0 0

0 Titles :

----------------------------------------------------
----------------------------------------------------

Many thanks in advance for looking into this!
Antje

Alexis Fri, 2015-07-24 11:57

Hi Antje,

The problem is that you didn't give the .mrc extension to the filename for the diagnostic output image.

Rather than ctff4_out_avg62, say ctff4_out_avg62.mrc, and I hope it'll work.

Alexis