[3dem] Model Refinement
Mohamed A Sayed
mohamed.sobhy at kaust.edu.sa
Wed Dec 5 02:55:42 PST 2018
Hello
I am new to cryo-em map reconstruction and want to ask for advice about improving the resolution of the final model. Please see below the steps that I applied.
I think my model current resolution is limited by my ability to perform proper refinement.
- Is there a way to improve the refinement and to check if this best possible final resolution obtained by the experimental data?
- At such resolution, is it worth performing molecular replacement since there is a pdb file already available from x-ray?
Thank you very much for your help.
I am using Relion 3.0 to performing Bayesian polishing, 3d auto-refine and generating mask on 83,000 particles, the final resolution came to be 4.27 A
mask paramters:
lowpass filter map 15 A
Initial binarisation threshold: 0.003
extend binary map this many pixels:0
add a soft-edge of this many pixels: 6
== Masking input maps ...
+ randomize phases beyond: 8.28125 Angstroms
== Dividing map by the MTF of the detector ...
+ mtf STAR-file: mtf2_k2_300kV.star
== Applying sqrt(2*FSC/(FSC+1)) weighting (as in Rosenthal & Henderson, 2003) ...
== Fitting straight line through Guinier plot to find B-factor ...
+ fit from resolution: 10
+ fit until resolution: 0
+ slope of fit: -43.0451
+ intercept of fit: -12.7354
+ correlation of fit: 0.908274
+ apply b-factor of: -172.18
== Low-pass filtering final map ...
+ filter frequency: 4.27419
== Writing output files ...
+ Processed map: PostProcess/job064/postprocess.mrc
+ Processed masked map: PostProcess/job064/postprocess_masked.mrc
+ Metadata file: PostProcess/job064/postprocess.star
+ FINAL RESOLUTION: 4.27419
I made another 3D classification on the shiny particles and selected one 3D class of 44,000 particles of the highest resolution performed 3d auto-refine, applied same mask and did post-processing my final resolution became 4.42 A
+ input mask: MaskCreate/job057/mask.mrc
== Masking input maps ...
+ randomize phases beyond: 8.0303 Angstroms
== Dividing map by the MTF of the detector ...
+ mtf STAR-file: mtf2_k2_300kV.star
== Applying sqrt(2*FSC/(FSC+1)) weighting (as in Rosenthal & Henderson, 2003) ...
== Fitting straight line through Guinier plot to find B-factor ...
+ fit from resolution: 10
+ fit until resolution: 0
+ slope of fit: -49.6022
+ intercept of fit: -12.9622
+ correlation of fit: 0.970846
+ apply b-factor of: -198.409
== Low-pass filtering final map ...
+ filter frequency: 4.41667
== Writing output files ...
+ Processed map: PostProcess/job078/postprocess.mrc
+ Processed masked map: PostProcess/job078/postprocess_masked.mrc
+ Metadata file: PostProcess/job078/postprocess.star
+ FINAL RESOLUTION: 4.41667
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