Extended Data Fig. 10: PolθH-DNA cryo-EM image analysis workflow.
From: Human polymerase θ helicase positions DNA microhomologies for double-strand break repair
Representative processing workflow described for DNA-bound PolθH. For 3D sorting jobs, representative classes are shown. In this scheme, the Ab initio model was utilized as a volume input for iterative heterogeneous refinement. Particles with apparent DNA density were subject to iterative 3D variability analysis with a mask encompassing the most heterogeneous regions to eliminate particles with no D5 density and poor DNA density. Particles were sorted into three groups based on DNA conformation (colored) and subject to non-uniform refinement to produce final reconstructions of PolθH in three different DNA pairing states, each above an angular distribution plot and a 3DFSC histogram with the map-to-model FSC overlaid. As some DNA density disappears upon map sharpening, a locally filtered map is also shown for each reconstruction.
