 |
 |
Hi Mike, We used them for all our ribosome structures (Amunts et al Science, Brown et al Science, Fernandez et al Cell, Wong et al eLife). We've recently also used it for gamma-secretase, but that hasn't been published yet. HTH, S > Hi Sjors, > > can you point to some publications that used these scripts? I’d like to > look at their results. > > Regards, > > mike > >> On May 20, 2015, at 4:49 AM, Sjors Scheres <[log in to unmask]> >> wrote: >> >> Dear all, >> Perhaps this is a good moment to highlight the link sent by Alan Brown >> below. It encompasses almost 2 years of experience in building and >> refining atomic models in high-resolution EM maps using COOT and REFMAC. >> At LMB, these scripts and instructions have become an important part of >> our structure determination pipeline. It may also be useful in your >> work. >> Best, >> Sjors >> >> >> On 05/20/2015 09:30 AM, Alan Brown wrote: >>> Hi Chris, >>> >>> We have a script (locres.csh), available from >>> http://www2.mrc-lmb.cam.ac.uk/groups/murshudov/content/em_fitting/em_fitting.html >>> that will calculate the local FSC for each chain in a folder of PDBs >>> using Relion for both the mask creation and the post-processing. >>> >>> Best wishes, >>> >>> Alan >>> >>> >>> >>> On 20 May 2015, at 08:36, Sjors Scheres <[log in to unmask]> >>> wrote: >>> >>>> Hi Chris, >>>> >>>> a. this is true: the 2 unfil.mrc will NOT be masked >>>> b. you'll calculate a final FSC in the postprocessing program. You can >>>> give any mask you like, and the randomized phase approach (see Chen et >>>> al >>>> Ultramicroscopy 2013) will correct for its effects on the FSC curve. >>>> If >>>> you let the mask cover only the best ordered bit of the map, it will >>>> be >>>> higher than when you include the more floppy bits. This gives an >>>> indication of local resolution, much like ResMap would. In a recenty >>>> accepted paper on the spliceosome tri-snRNP we did this for many small >>>> regions, and got very similar results as resmap. >>>> Make sure your masks are always with a soft-edge, so their effects on >>>> the >>>> FSC are not very large. It doesn't matter which program you use to >>>> make >>>> the masks. >>>> HTH, >>>> Sjors >>>> >>>> >>>>> Sjors and users- >>>>> >>>>> I am using a mask during refinements in Relion, which eliminates the >>>>> small >>>>> ribosomal subunit, thus, keeping its multiple conformations from >>>>> affecting >>>>> alignments driven by the lsu. >>>>> I created this mask from a volume "made" in Chimera and then used >>>>> relion_mask_create. >>>>> >>>>> My question is two-fold: >>>>> >>>>> a. The 3d half volumes made at each itr are masked by this volume, >>>>> but I >>>>> presume that the final 3d volume >>>>> that is made when Relion converges will not be masked by this volume, >>>>> is >>>>> this correct? >>>>> >>>>> b. Given that part a above is true, then I presume that the final fsc >>>>> calculated automatically in Relion will reflect the >>>>> properties of the entire 3D volume, including the region that had >>>>> been >>>>> masked out during >>>>> refinement, which may lower the overall resolution. >>>>> >>>>> While I can think of a number of ways to make a mask, what is the >>>>> appropriate or best way to make >>>>> a mask and the use Relion to calculate an fsc for the good region of >>>>> the >>>>> map. >>>>> >>>>> Of course ResMap can be used as a guide, but getting an fsc for the >>>>> good >>>>> region would be useful and >>>>> in my reading its not clear how this is done. >>>>> >>>>> cheers, >>>>> >>>>> C Akey >>>>> >>>> >>>> -- >>>> Sjors Scheres >>>> MRC Laboratory of Molecular Biology >>>> Francis Crick Avenue, Cambridge Biomedical Campus >>>> Cambridge CB2 0QH, U.K. >>>> tel: +44 (0)1223 267061 >>>> http://www2.mrc-lmb.cam.ac.uk/groups/scheres >>> >> >> -- >> Sjors Scheres >> MRC Laboratory of Molecular Biology >> Francis Crick Avenue, Cambridge Biomedical Campus >> Cambridge CB2 0QH, U.K. >> tel: +44 (0)1223 267061 >> http://www2.mrc-lmb.cam.ac.uk/groups/scheres > -- Sjors Scheres MRC Laboratory of Molecular Biology Francis Crick Avenue, Cambridge Biomedical Campus Cambridge CB2 0QH, U.K. tel: +44 (0)1223 267061 http://www2.mrc-lmb.cam.ac.uk/groups/scheres