If you have a preliminary solution, you can use the Fcalcs (e.g. from Phaser or Refmac) as a reference in Aimless, and get an analysis of the agreement between the scaled unmerged data and the reference as a function of Batch (~= Image number). This may indicate that some parts of the the data agree better with your model than others. Of course there is an inherent bias in this analysis, but it may still be informative.
Phil
On 19 Jul 2015, at 17:15, Christopher Barnes <[log in to unmask]> wrote:
> Harry- Yes we have used pointless when merging multiple sets, and we always come back with p212121.
>
> Tim - Yes i am confident that the values are not a by-product of generally little data per wedge, as some wedges containing more data (because of thicker regions of the crystal) produce the I/sigI or Isa values that I mentioned in my previous message.
>
> Kay - As for the indexing ambiguity, two of the axes are pretty close (within 5 Angstroms of each other), and so we do sometimes see programs index in higher symmetry (P4) spacegroup. However, I have collected multiple low dose datasets from single crystals to ~4.4 Angstrom (at a BM-line, so no translation), where you can clearly see that P212121 is most likely the right spacegroup (ISa for P212121 > 30 whereas ISa for P4 related spacegroups is never greater than 2, also determined using pointless as well). In addition, these crystals are composed of a multi-protein complex, where 75% of the components are structurally known; therefore, molecular replacement in p212121 gives a clear solution with no symm-related clashes, and initial rigid/bgroup refinement gives Rwork/Rfree values of 0.29/0.34. Maps also show a good fit for the known parts of the model, with clear electron density in the difference map for the novel components.
>
> As for the radiation damage, I wonder why this would cause such a problem, since we are only losing the high res reflections? So while we have datasets to 4.5 Angstrom, at that resolution there is no way to trace the novel components (se-mets are sparse, and generally reside in predicted disordered loop regions which would not be much help for tracing). So translating after a few degrees is the only way to maintain the high intensity reflections we need at the higher resolution (~3.5 Angstrom, which already is pretty weak). However, reflections less than 4.2 stay consistent (at least visually), so why would radiation damage between the last frame from one wedge and first frame of the next wedge cause such a dramatic drop even for the low res data, and is there any way I can correct for this?
>
> Thanks,
> Christopher
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