B-factor value is too high in Non uniform refinement

3D Refinement
1

Hi,
I am facing problem related to high b-factor (800-1100) while running Non -Uniform refinement in CryoSPARC version 3.2.
Previously, same dataset I processed in version 2.4. At that time,I didn’t face this problem.
Can someone please help me to figure out what I should do to resolve this issue.

Thanks,

Regards,
Priyanka

2

The used B-factor doesn’t affect your processing, so you shouldn’t worry about the automatically applied one.

You can input the output volume from your refinement into a “Sharpening Tools” job in the utilities section in order to apply the B-factor of your choosing to your map.

You can also download the map and input it into another program to “optimally” auto-sharpen the map.

3

Hi,
Thanks for your reply.
Can you suggest what range of b-factor value I can try?
What is the difference between Non-Uniform refinement New and Non-Uniform refinement legacy in version 3.2 ?

Thanks,

Regards,
Priyanka

4

The appropriate b-factor to use is dataset and resolution dependent, so I can’t really give you one to use; you should try a few and see which one fits you best (or you can use another program to calculate the most optimal one(s)).

You can read all about non-uniform refinement here:
https://guide.cryosparc.com/processing-data/all-job-types-in-cryosparc/3d-refinement/job-non-uniform-refinement-new

and here

https://www.nature.com/articles/s41592-020-00990-8

Best,
Armando