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Rescaled refined particles extraction error #971

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janti001 opened this issue May 5, 2023 · 4 comments
Closed

Rescaled refined particles extraction error #971

janti001 opened this issue May 5, 2023 · 4 comments

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@janti001
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janti001 commented May 5, 2023

I have been trying to perform a particle extraction job with refined particles that I have obtained from a round of 3D classification and selection jobs. The particles went through several rounds of 2D classification (~200k particles) (0.651 A/pix rescaled to 1.302A/pix) before being put through a 3D classification and selection job (as mentioned above), (~40k particles). A previous refined particle extraction using a different, not as good, 3D classification job output, seemed to work fine (~80 000 particles, 1.302A/pix) (extracted the same amount of particles that were inputed into the extraction job). However, with my current, improved 3D classifications rescaled refined particles extraction is completely off. I have
~40 000 particles that require extracting, however, I end up extracting, after several attempts, numerous amounts of particles, >100 000 which makes no sense, how can it extract more particles than is being inputted?

The only difference between the two 3D classification jobs is the angular sampling interval (3.7 to 1.8), all inputs and other parameters are identical so I am not sure what may be causing this issue. Both of the extraction jobs are identical, with the only difference being the input coordinates. I checked the particles.star file from the inputs into the successful and failed extraction jobs seem fine.

Particles.star file input for successful extraction job:

version 30001

data_optics

loop_
_rlnOpticsGroupName #1
_rlnOpticsGroup #2
_rlnMtfFileName #3
_rlnMicrographOriginalPixelSize #4
_rlnVoltage #5
_rlnSphericalAberration #6
_rlnAmplitudeContrast #7
_rlnImagePixelSize #8
_rlnImageSize #9
_rlnImageDimensionality #10
opticsGroup1 1 processing/K3_CDS_MTF.star 0.651000 300.000000 2.700000 0.100000 1.302000 280 2

version 30001

data_particles

loop_
_rlnCoordinateX #1
_rlnCoordinateY #2
_rlnHelicalTubeID #3
_rlnAngleTiltPrior #4
_rlnAnglePsiPrior #5
_rlnHelicalTrackLengthAngst #6
_rlnAnglePsiFlipRatio #7
_rlnImageName #8
_rlnMicrographName #9
_rlnOpticsGroup #10
_rlnCtfMaxResolution #11
_rlnCtfFigureOfMerit #12
_rlnDefocusU #13
_rlnDefocusV #14
_rlnDefocusAngle #15
_rlnCtfBfactor #16
_rlnCtfScalefactor #17
_rlnPhaseShift #18
_rlnGroupNumber #19
_rlnAngleRot #20
_rlnAngleTilt #21
_rlnAnglePsi #22
_rlnOriginXAngst #23
_rlnOriginYAngst #24
_rlnClassNumber #25
_rlnNormCorrection #26
_rlnLogLikeliContribution #27
_rlnMaxValueProbDistribution #28
_rlnNrOfSignificantSamples #29
4890.992676 2955.164795 7 57.978448 32.433463 0.000000 0.500000 000076@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 -76.40540 57.978448 32.433463 6.060745 13.820203 1 0.660921 68815.409690 0.035791 3389
5007.459473 2915.802490 7 80.845617 -149.88697 80.033046 0.500000 000078@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 34.099722 80.845617 -149.88697 12.838472 17.213922 1 0.660931 68814.702321 0.079008 3576
3572.893799 817.126038 4 98.833703 64.316189 229.189341 0.500000 000045@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 -11.22739 98.833703 64.316189 9.579702 7.159340 1 0.662410 68657.378307 0.084045 2926
3609.261475 771.060486 4 76.094959 64.883374 267.397234 0.500000 000046@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 -174.79278 76.094959 64.883374 16.952163 10.791815 1 0.662222 68713.940148 0.122545 4000
3645.628906 724.994934 4 105.154772 -115.11020 305.605029 0.500000 000047@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 -174.22237 105.154772 -115.11020 7.787571 0.928852 1 0.663098 68726.543470 0.059270 4263
3378.062988 1035.947021 4 84.763720 -129.06954 38.207755 0.500000 000040@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 -28.31664 84.763720 -129.06954 -4.91085 -7.53158 1 0.662506 68747.805909 0.226378 2308
945.543579 3620.742676 3 96.406743 -133.29111 113.244175 0.500000 000037@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 -112.05939 96.406743 -133.29111 -6.39735 -8.46830 1 0.663914 68774.765300 0.146673 1897
3419.394531 994.277466 4 86.880637 -129.18542 76.415678 0.500000 000041@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 101.532802 86.880637 -129.18542 -1.73642 -5.24424 1 0.663924 68772.281414 0.224854 3379
3336.731689 1077.616455 4 99.464271 -128.53919 0.000000 0.500000 000039@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 -33.27902 99.464271 -128.53919 -1.92042 1.294244 1 0.662909 68772.744494 0.014288 5826
1249.561401 2724.541016 1 99.154383 74.886970 383.032344 0.500000 000011@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 -145.90028 99.154383 74.886970 -26.16695 -13.54133 1 0.656882 68848.974358 0.017431 3462
1214.846802 2897.606445 1 121.972455 73.587222 268.122552 0.500000 000008@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 42.031034 121.972455 73.587222 -11.98867 -6.32118 1 0.661311 68845.805726 0.019260 2727
5058.491699 2882.923828 7 81.166297 -169.31619 119.553060 0.500000 000079@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 168.772606 81.166297 -169.31619 -1.08141 -4.54644 1 0.664729 68751.434817 0.016074 3220
5026.923340 3578.219238 5 104.823226 61.503856 39.183796 0.500000 000053@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 151.307529 104.823226 61.503856 11.261722 4.825756 1 0.663443 68645.168395 0.062944 3318
5106.554199 2844.603271 7 99.776596 51.455180 159.569543 0.500000 000080@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 -157.24881 99.776596 51.455180 1.901165 6.984574 1 0.663871 68776.538436 0.012795 3011
4426.540039 3612.254150 10 95.560685 134.727565 0.000000 0.500000 000099@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 38.752118 95.560685 134.727565 -17.01442 20.711267 1 0.659611 68760.047291 0.072697 2645
2948.490967 1572.234375 2 93.110688 50.817276 331.431254 0.500000 000028@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 67.033954 93.110688 50.817276 -11.43557 -10.92860 1 0.657845 68935.697267 0.049838 2880
5154.616699 2806.282715 7 108.979138 40.667923 199.586026 0.500000 000081@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 -0.30837 108.979138 40.667923 -4.90437 -6.95466 1 0.664633 68761.484169 0.025423 3187
3742.845215 578.261902 4 86.880637 58.314579 420.198360 0.500000 000050@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 101.532802 86.880637 58.314579 5.147497 3.950607 1 0.662139 68665.568013 0.017595 3243
2912.017090 1615.473389 2 93.143274 51.527500 294.605386 0.500000 000027@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 117.903435 93.143274 51.527500 -8.58981 -5.84743 1 0.660328 68895.631733 0.034408 4230

Particles.star file for erroneous refined particle extraction job:

version 30001

data_optics

loop_
_rlnOpticsGroupName #1
_rlnOpticsGroup #2
_rlnMtfFileName #3
_rlnMicrographOriginalPixelSize #4
_rlnVoltage #5
_rlnSphericalAberration #6
_rlnAmplitudeContrast #7
_rlnImagePixelSize #8
_rlnImageSize #9
_rlnImageDimensionality #10
opticsGroup1 1 processing/K3_CDS_MTF.star 0.651000 300.000000 2.700000 0.100000 1.302000 280 2

version 30001

data_particles

loop_
_rlnCoordinateX #1
_rlnCoordinateY #2
_rlnHelicalTubeID #3
_rlnAngleTiltPrior #4
_rlnAnglePsiPrior #5
_rlnHelicalTrackLengthAngst #6
_rlnAnglePsiFlipRatio #7
_rlnImageName #8
_rlnMicrographName #9
_rlnOpticsGroup #10
_rlnCtfMaxResolution #11
_rlnCtfFigureOfMerit #12
_rlnDefocusU #13
_rlnDefocusV #14
_rlnDefocusAngle #15
_rlnCtfBfactor #16
_rlnCtfScalefactor #17
_rlnPhaseShift #18
_rlnGroupNumber #19
_rlnAngleRot #20
_rlnAngleTilt #21
_rlnAnglePsi #22
_rlnOriginXAngst #23
_rlnOriginYAngst #24
_rlnClassNumber #25
_rlnNormCorrection #26
_rlnLogLikeliContribution #27
_rlnMaxValueProbDistribution #28
_rlnNrOfSignificantSamples #29
4890.992676 2955.164795 7 109.504012 -147.22068 0.000000 0.500000 000076@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 136.304551 109.504012 -147.22068 5.819338 12.297787 1 0.660179 1.528419e+05 0.076513 23815
5007.459473 2915.802490 7 92.432658 -150.94529 80.033046 0.500000 000078@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 170.065291 92.432658 -150.94529 13.115287 22.145289 1 0.659647 1.527851e+05 0.279027 13781
3572.893799 817.126038 4 86.456626 -126.58393 229.189341 0.500000 000045@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 151.785701 86.456626 -126.58393 11.135026 5.825915 1 0.662568 1.528967e+05 0.024489 14663
1226.418335 2839.917969 1 97.136114 -111.55466 306.425816 0.500000 000009@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 -9.93340 97.136114 -111.55466 -14.83590 -6.65319 1 0.660740 1.530622e+05 0.053837 10763
3609.261475 771.060486 4 90.009894 -115.35672 267.397234 0.500000 000046@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 102.565615 90.009894 -115.35672 18.124935 10.641301 1 0.662039 1.528368e+05 0.265703 19205
914.570740 1679.871216 9 80.364490 90.918907 184.141190 0.500000 000097@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 23.818725 80.364490 90.918907 3.228998 1.329816 1 0.663492 1.530035e+05 0.033785 29716
3336.731689 1077.616455 4 86.457628 47.793056 0.000000 0.500000 000039@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 154.598077 86.457628 47.793056 -10.13416 -7.08314 1 0.660643 1.529394e+05 0.112086 14313
1249.561401 2724.541016 1 99.553607 70.333193 383.032344 0.500000 000011@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 -26.80594 99.553607 70.333193 -25.52053 -8.94296 1 0.657866 1.530753e+05 0.064705 14591
4512.474121 480.310608 12 105.680258 -113.46908 41.323506 0.500000 000104@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 42.088325 105.680258 -113.46908 -11.90513 -6.05038 1 0.658826 1.530852e+05 0.325642 22536
5026.923340 3578.219238 5 76.791968 -141.54059 39.183796 0.500000 000053@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 -151.97064 76.791968 -141.54059 -13.61916 -20.25395 1 0.657661 1.529149e+05 0.041430 15940
5106.554199 2844.603271 7 109.516191 -152.81842 159.569543 0.500000 000080@Extract/job018/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrcs MotionCorr/job002/Movies/GridSquare_2797388/Data/FoilHole_4447588_Data_4432512_4432514_20230205_134403_fractions.mrc 1 4.289456 9.650000e-04 33766.281250 33698.328125 -49.30104 0.000000 1.000000 0.000000 1 172.876143 109.516191 -152.81842 -10.32171 -22.08009 1 0.660807 1.527200e+05 0.158612 23951
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956.420349 1484.688354 5 79.231965 83.461183 150.382158 0.500000 000048@Extract/job018/Movies/GridSquare_2797388/Data

@biochem-fan
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Can you inspect the output STAR file and find which line in the input STAR file generated multiple output particles?

@janti001
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janti001 commented May 5, 2023

I have checked and there seems to be no duplication, the error seems to be that the program is extracting particles from micrographs it does not have co-ordinated for? The input partices.star file has co-ordinates for a batch of micrographs exclusively named gridsquare_2797388, but in the output file it is searching through batch 2797388 and 2797487? I checked that the first 40k lines are for 2797388, which is the correct amount of particles, and then it starts to extract micrographs that it has no coordinates for?

@janti001
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janti001 commented May 5, 2023

For context as well, The extraction job was submitted to a queue system, at the time of submission, all nodes were occupied and for some reason, the job ran instantaneously and gave the particle.star file with too many particles? It then got through to the queuing system and then processed but gave the same output?

@janti001
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janti001 commented May 5, 2023

Resubmitted the job to queue and all seems to be working now, no problems. Likely was an issue with the queueing system. Thank you.

@janti001 janti001 closed this as completed May 5, 2023
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