Hi,
Probably your B factor refinement was not successful.
(It does not necessarily work nicely as I explained in the previous mail)
Please use STAR files before B factor refinement or
remove rlnCtfBfactor and rlnCtfScalefactor columns.
Best regards,
Takanori Nakane
> Hi,
>
> ??? I see a problem that is possibly related to the one just reported
> by several people: Refinement in relion-3.1-beta runs nicely but upon
> combining the two half maps it reports a resolution of 0.871938 A, which
> is obviously impossible. Nevertheless, postprocessing works fine giving
> me realistic resolution but the FSC curve is messed up similarly as
> reported by Minglei; it goes up again at the right side. I noticed that
> in the last iteration several warnings were issued like:
>
> ? 112590 in group 13; Are your groups large enough? Or is the reference
> on the correct greyscale?
> 2.19/2.19 hrs
> ............................................................~~(,_,">
>
> I also noticed that some values of? _rlnLogLikeliContribution #23 were
> negative in the final run_data.star!
>
> Nevertheless, the final map looks good!
>
> best, Dieter
>
> ------------------------------------------------------------------------
> Dieter Blaas
> Max Perutz Laboratories
> Medical University of Vienna,
> Inst. Med. Biochem., Vienna Biocenter (VBC),
> Dr. Bohr Gasse 9/3,
> A-1030 Vienna, Austria,
> Tel: 0043 1 4277 61630,
> Mobile: 0043 699 1942 1659
> Fax: 0043 1 4277 9616,
> e-mail: [log in to unmask]
> ------------------------------------------------------------------------
>
> Am 04.12.2019 um 13:53 schrieb Takanori Nakane:
>> Hi,
>>
>> Indeed B factor refinement is still new and we don't know when
>> it is beneficial. I recommend you to leave it OFF during processing
>> and only try it at the very end to see if it improves the resolution.
>>
>> You can also try these options in relion_ctf_refine to avoid
>> excessive B factors.
>>
>> --bfac_min_B (-30) : Minimal allowed B-factor
>> --bfac_max_B (300) : Maximal allowed B-factor
>>
>> For example, you can make the range smaller (say, 0 to 150).
>>
>> Best regards,
>>
>> Takanori Nakane
>>
>>> FYI I've seen a similar effect at FSC high-frequencies when using
>>> B-factor refinement in relion v3.1
>>>
>>> Cheers,
>>> Joe
>>>
>>> On 03/12/2019, 21:08, Minglei Zhao wrote:
>>>> OK. Will report back.
>>>>
>>>> Thanks!
>>>>
>>>> Minglei
>>>>
>>>>> On Dec 3, 2019, at 2:50 PM, Takanori Nakane
>>>>> <[log in to unmask]>
>>>>> wrote:
>>>>>
>>>>> Hi,
>>>>>
>>>>> Interesting.
>>>>>
>>>>>> After CtfRefine (first beam tilt, then anisotropic magnification,
>>>>>> then
>>>>>> defocus)
>>>>> Can you run Refine3D on outputs from each step to see which one was
>>>>> harmful?
>>>>>
>>>>> Best regards,
>>>>>
>>>>> Takanori Nakane
>>>>>
>>>>> On 2019/12/03 20:33, Minglei Zhao wrote:
>>>>>> Just double checked, no intermediate half maps were used in any post
>>>>>> processing.
>>>>>> Minglei
>>>>>>> On Dec 3, 2019, at 2:30 PM, Takanori Nakane
>>>>>>> <[log in to unmask]> wrote:
>>>>>>>
>>>>>>> Hi,
>>>>>>>
>>>>>>> First of all, is "Relion 3.0 beta" typo of
>>>>>>> RELION 3.1 beta? Otherwise, you should use at least RELION 3.0.8.
>>>>>>>
>>>>>>>> the high frequency signal is abnormal (why?), but the curve still
>>>>>>>> looks
>>>>>>> Probably you used half maps from intermediate iterations in
>>>>>>> PostProcess.
>>>>>>> Please use the converged half maps and repeat subsequent steps.
>>>>>>>
>>>>>>> Best regards,
>>>>>>>
>>>>>>> Takanori Nakane
>>>>>>>
>>>>>>> On 2019/12/03 20:25, Minglei Zhao wrote:
>>>>>>>> Dear Microscopists,
>>>>>>>> I encountered a weird behavior of FSC for an apoferritin dataset
>>>>>>>> processed using Relion 3.0 beta:
>>>>>>>> After 3D refinement and post process, everything looks good:
>>>>>>>> After CtfRefine (first beam tilt, then anisotropic magnification,
>>>>>>>> then defocus), refined again, and post process, the high frequency
>>>>>>>> signal is abnormal (why?), but the curve still looks OK before
>>>>>>>> 0.7(1/A). Estimated resolution improved a little bit:
>>>>>>>> After Bayesian Polishing, refined again using mask and
>>>>>>>> solvent-flattened FSC, the estimated resolution is 1.9 A, FSC from
>>>>>>>> Refine3D job looks fine:
>>>>>>>> However, using the same mask in the previous refinement to filter
>>>>>>>> and sharpen the map, the FSC curve is messed up:
>>>>>>>> Any obvious things I did wrong?
>>>>>>>> Thanks for your attention.
>>>>>>>> Minglei
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