Create a workflow for cryo-electron microscopy single-particle analysis: (1) protein sample vitrification on holey carbon grid (Vitrobot), (2) cryo-EM data collection on 300kV microscope (Titan Krios) with K3 detector, (3) movie frame alignment and CTF estimation (MotionCor2, CTFFIND4), (4) automated particle picking (crYOLO/TOPAZ), (5) 2D classification and class selection, (6) ab initio 3D reconstruction, (7) 3D refinement with gold-standard FSC, (8) CTF refinement and Bayesian polishing, (9) local resolution estimation, (10) atomic model building (Coot) and real-space refinement (Phenix). Show resolution milestones.
