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turonova / novaCTF

3D-CTF correction for cryo-electron tomography
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How to do the alignment after doing ctfcorrection? #19

Open XinYou-Tshua opened 3 years ago

XinYou-Tshua commented 3 years ago

Hi,

I am trying to do the tomograms reconstruction by Novactf. However, some troubles appeared. My workflow was described as below: 1). I use CTFFIND4.1.8 to estimate the defocus of original stack file (named tomo.st), generating a ' diagnostic_output.txt ' file (renamed tomo_defocus.txt).

2). I input tomo.st, tomo.tlt and use tomo_defocus.txt as the DefocuseFile. Then I run the script: novaCTF -Algorithm defocus -InputProjections tomo.st -FULLIMAGE 3838,3710 -THICKNESS 108 -TILTFILE tomo.tlt -SHIFT 0.0,0.0 -CorrectionType phaseflip -DefocusFileFormat ctffind4 -CorrectAstigmatism 1 -DefocusFile tomo_defocus.txt -PixelSize 0.3421 -DefocusStep 15 This generated a lot of tomodefocus.txt# files.

3). Then I do the ctfcorrection by script: for i in $(ls txt_) do novaCTF -Algorithm ctfCorrection -InputProjections tomo.st -OutputFile $i.st -DefocusFile $i -TILTFILE tomo.tlt -CorrectionType phaseflip -DefocusFileFormat ctffind4 -CorrectAstigmatism 1 -PixelSize 0.3421 -AmplitudeContrast 0.07 -Cs 2.7 -Volt 300 done This generated a lot of stack files.

My question is how should I do the alignment and clip in the next step use these stack files? I just used ETOMO-GUI to do the fine alignment, however, there are too many stack files in the last step, so I have no idea to do that. Would you mind give me a detailed steps? I am looking forward to your reply ASAP.

Best, Xin

zehrea commented 2 years ago

Hi Xin,

Did you get an answer? I have the same question as you are.

Thanks, Elena

XinYou-Tshua commented 2 years ago

Hi zehera,

You can do it by 'newstack' command in IMOD. And a .xf file is necessary. This file is generated by fine alignment in ETOMO gui. like this: newstack -x .xf -bin 4 -in .st -ou ***.ali

Hope it can help you. Best, Xin

zehrea commented 2 years ago

Hi Xin,

Thank you for your response! I am following your advice above but I am getting error

newstack -x 2021-10-19_tomo001.xf -bin 3 -in .st -ou .ali ERROR: NEWSTACK - You must specify number of sections to write to each output file

How do you get around this?

Thanks, Elena

XinYou-Tshua commented 2 years ago

Hi Elena.

Indeed, I did not get this error yet. However, please check you input file one by one. And could you provide your original command? I do not sure your '*.st' file is raw tilt stack.

Best, Xin