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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