Error from particle re-extraction

Troubleshooting
1

hi everyone,

I exported my particles from cryosparc (128 px at 1.059 A/px) to Relion to do 3D classification and 3D Refinement. After 3D refinement I tried to re-extract in a larger box size From 128px to 256px and encountered this error:

Warning: There are only 9 particles in micrograph J3/motioncorrected/10003260799687496747_stack_04055_X+0Y+0-0_patch_aligned_doseweighted.mrc. Consider joining multiple micrographs into one group. 
 micrograph x,y,z,n-size= 5760 , 4092 , 1 , 1
 particle position= 3739 , 5153
in: /packages/RELION/github/version/3.1.1/src/preprocessing.cpp, line 808
ERROR: 
Preprocessing::extractParticlesFromOneFrame ERROR: particle4 lies completely outside micrograph J3/motioncorrected/10003260799687496747_stack_04055_X+0Y+0-0_patch_aligned_doseweighted.mrc
=== Backtrace  ===
/opt/relion/3.1.1/bin/relion_preprocess(_ZN11RelionErrorC1ERKNSt7__cxx1112basic_stringIcSt11char_traitsIcESaIcEEES7_l+0x67) [0x564ad9d78c77]
/opt/relion/3.1.1/bin/relion_preprocess(+0xa81da) [0x564ad9d9e1da]
/opt/relion/3.1.1/bin/relion_preprocess(_ZN13Preprocessing31extractParticlesFromFieldOfViewE8FileNamel+0x6b0) [0x564ad9d9f030]
/opt/relion/3.1.1/bin/relion_preprocess(_ZN13Preprocessing19runExtractParticlesEv+0x161) [0x564ad9da0671]
/opt/relion/3.1.1/bin/relion_preprocess(_ZN13Preprocessing3runEv+0x12) [0x564ad9da12c2]
/opt/relion/3.1.1/bin/relion_preprocess(main+0x3c) [0x564ad9d6db2c]
/lib/x86_64-linux-gnu/libc.so.6(__libc_start_main+0xeb) [0x7ff8ee53809b]
/opt/relion/3.1.1/bin/relion_preprocess(_start+0x2a) [0x564ad9d70e3a]
==================
ERROR: 
Preprocessing::extractParticlesFromOneFrame ERROR: particle4 lies completely outside micrograph J3/motioncorrected/10003260799687496747_stack_04055_X+0Y+0-0_patch_aligned_doseweighted.mrc

I swapped XY with csparc2star.py and repeated Class3D but the particles look weird after swapping xy which makes me believe that the problem is not that.
or I’m doing something wrong.

Any insight on what might be the problem would be very much appreciated.

thank you

2

Hi @Proteino

Did you ever figure out what was going on here? I’m experiencing a similar issue with one of my datasets, and I’m totally stumped.

I’ve tried --inverty, --invertx, --invertx --inverty, --swapxy, etc., basically every iteration possible, and still no luck when I try to re-extract the cs-derived particles in relion. I get exactly the same error (ie, particle# lies completely outside micrograph XYZ).

I’ve successfully deployed csparc2star.py on 3 different cs-derived particle sets (thx so much @DanielAsarnow!!) from other datasets without fail, and can’t seem to figure out what I did wrong with this one.

pls halp! :pray:t4:

3

@Cryo_EM_Jonny double check the pixel sizes?