As my sample is quite heterogeneous, I tried 3DVA for resolving confomation heterogeneity.
My question is about the usage of ‘Clustered Particles and associated Volume’, generated by 3DVA->3DVA-display clustered mode. The 3DVA was performed with 270K particles and 3-variability components; and displayed by 3DVA-display cluster mode with 20 clusteres that gave 20 particles-clustes with associated volumes. Could I use the particle-cluster and associated volume directly for homogenous refinement? I already tried this and it gave refined-maps (3.5-3.8 A with approx. 15K particles) showing resonable features of side-chains. Is there any chance of introducing bias with this approach?
Yes, this is an acceptable way to get your final refinement and discrete particle set, as long as you gave a decent lowpass filter for the refinement (or not mess with the default value). You may also try clustering fewer components (eg try component 0 and component 0, rather than all 3 at once) by removing and replacing items in the lower level input.
thank you for your reply and suggeting of using individual component for clustering. I tried to use individual component by removing particles and volume of other components from lower level input. The job failed with following error: IndexError: index 1 is out of bounds for axis 1 with size 1.
Yes - you have to replace all the existing components with the desired component. So if you have four components in the initial 3D-VA job, you will need to provide four copies of e.g. component 1, for example.
I ran 3DVA with 3 components, then ran 3DVA analysis cluster mode with just 2 copies of the same component. Is this ok? I never tried with 3 copies of the same component.
should be fine if it works! In my hands if I don’t have the same number of components as the input cryosparc throws an error. @team it would be great if clustering using a subset of components was “officially” supported, I have found it very useful for isolating low occupancy states
@olibclarke This is now officially supported cryoSPARC 3.3! Thanks for the nudge . The 3DVA Display job includes an advanced parameter called Only use these components which can select specific components to use for all modes.
. The 3DVA Display job includes an advanced parameter called