Scalepack help
Zbyszek Otwinowski
zbyszek at mix.swmed.edu
Wed Oct 9 20:05:14 EST 1996
Tim Allison wrote:
>
> I'm having trouble in Scalepack and hope someone can help.
>
> My limited experience has been that, at least in initial rounds,
> before tinkering with error values, the value of Chi^2 increases with
> increasing resolution. That is, if I set "estimated error" to 0.04
> and "error scale factor" to 1.3, Chi^2 may be 1 or 2 for low
> resolution shells and increase up to 4,5, maybe 10 for the highest
> resolution shell.
This is sign that something wrong is in the experiment or in data
reduction. I would expect Chi^2 values to be in the range 0.8-2.0
when experiment is done well and the crystal is single, does not
change due to radiation damage etc.
>
> I'm currently reducing and scaling SeMet MAD data from the NSLS (Fuji
> scanner). Instead of the situation I describe above, I'm finding that
> my highest resolution shell (1.66-1.6A) has Chi^2 of around 1.0 and my low
> resolution shells are around 3 or 4. I've tried an array of error
> scale factors, estimated errors, and rejection probabilities but can't
> seem to lower the Chi^2 of the low res shells to below 2.0.
>
Use option "scale anomalous". Otherwise anomalous signal contributes
to Chi^2. Without scale anomalous, one should expect _large_ Chi^2 at
low resolution in MAD experiment. Otherwise there there is no measurable
anomalous signal.
> Included below are sample scalepack inputs.
>
> So, my question is this: Is my experience simply too limited; is this
> a normal phenomenon? Is this indicative of something related to my
> use of denzo? Overloads? Mis-indexing? Spot area? Anomalous signal?
>
> I've scaled with "anomalous" off, on and with "scale anomalous" on;
> all give the same result.
>
> Additionally, what are reasonable values for Chi^2? Do most people
> try to refine them down to 1.0 or do they not worry about it and
> accept higher values?
>
> Thanks for any help and/or advice you can offer,
> Tim
>
> --------------------------------------------------------------
> Tim Allison Dept of Biochemistry
> tja2f at dayhoff.med.virginia.edu University of Virginia
> --------------------------------------------------------------
>
> -----------
> se1a.scpk:
> /net/rule1/tja2f/denzo.1.5.4/scalepack <<stopinp > se1a.splog
> number of zones 10
> estimated error 0.06 0.06 0.06 0.06 0.06 0.06 0.06 0.06 0.06 0.06
> error scale factor 1.5
> rejection probability 0.0001
> ignore overloads
> [@reject]
> write rejection file 0.9
> space group P21
> resolution 100 1.6
> sigma cutoff -3.0
> reference batch 01
> add partials 01 to 80
> postrefine 10
> fit crystal a* 01 to 80
> fit crystal b* 01 to 80
> fit crystal c* 01 to 80
> fit crystal beta* 01 to 80
> fit batch rotx 01 to 80
> fit batch roty 01 to 80
> fit crystal mosaicity 1 to 80
> anomalous
> format DENZO_york1
> sector 01 to 80
> file 'se1_a0##.y2'
> output file 'se1a.spout'
> stopinp
> echo 'Scalepack DONE'
>
> #I then rerun with @reject uncommented and "write rejection file 0.5"
>
> ----------
> typical output (using error scale factor 1.3 and estimated error of
> 0.06 for all shells:
>
> Shell Summary of observation redundancies by shells:
> Lower Upper No. of reflections with given No. of observations
> limit limit 0 1 2 3 4 5-6 7-8 9-12 13-19 >19 total
> 100.00 3.45 23 8 226 494 646 210 10 0 0 0 1594
> 3.45 2.74 10 0 143 449 712 251 5 0 0 0 1560
> 2.74 2.39 5 1 126 321 823 286 0 0 0 0 1557
> 2.39 2.17 2 1 104 271 866 309 0 0 0 0 1551
> 2.17 2.02 3 1 111 255 871 331 0 0 0 0 1569
> 2.02 1.90 4 2 93 221 880 355 0 0 0 0 1551
> 1.90 1.80 2 2 99 208 876 369 0 0 0 0 1554
> 1.80 1.72 3 22 179 506 591 245 0 0 0 0 1543
> 1.72 1.66 0 49 291 668 395 172 0 0 0 0 1575
> 1.66 1.60 30 204 415 487 292 109 0 0 0 0 1507
> All hkl 82 290 1787 3880 6952 2637 15 0 0 0 15561
>
> Shell Summary of observation redundancies:
> Lower Upper % of reflections with given No. of observations
> limit limit 0 1 2 3 4 5-6 7-8 9-12 13-19 >19 total
> 100.00 3.45 1.4 0.5 14.0 30.6 40.0 13.0 0.6 0.0 0.0 0.0 98.6
> 3.45 2.74 0.6 0.0 9.1 28.6 45.4 16.0 0.3 0.0 0.0 0.0 99.4
> 2.74 2.39 0.3 0.1 8.1 20.6 52.7 18.3 0.0 0.0 0.0 0.0 99.7
> 2.39 2.17 0.1 0.1 6.7 17.5 55.8 19.9 0.0 0.0 0.0 0.0 99.9
> 2.17 2.02 0.2 0.1 7.1 16.2 55.4 21.1 0.0 0.0 0.0 0.0 99.8
> 2.02 1.90 0.3 0.1 6.0 14.2 56.6 22.8 0.0 0.0 0.0 0.0 99.7
> 1.90 1.80 0.1 0.1 6.4 13.4 56.3 23.7 0.0 0.0 0.0 0.0 99.9
> 1.80 1.72 0.2 1.4 11.6 32.7 38.2 15.8 0.0 0.0 0.0 0.0 99.8
> 1.72 1.66 0.0 3.1 18.5 42.4 25.1 10.9 0.0 0.0 0.0 0.0 100.0
> 1.66 1.60 2.0 13.3 27.0 31.7 19.0 7.1 0.0 0.0 0.0 0.0 98.0
> All hkl 0.5 1.9 11.4 24.8 44.4 16.9 0.1 0.0 0.0 0.0 99.5
>
> Shell I/Sigma in resolution shells:
> Lower Upper No. of reflections with I / Sigma less than
> limit limit 0 1 2 3 5 10 20 >20 total
> 100.00 3.45 1 3 8 13 22 48 130 1464 1594
> 3.45 2.74 3 10 23 33 58 107 309 1251 1560
> 2.74 2.39 7 19 37 60 111 224 589 968 1557
> 2.39 2.17 8 30 67 119 184 375 830 721 1551
> 2.17 2.02 12 56 119 197 296 538 1004 565 1569
> 2.02 1.90 37 120 214 312 460 743 1218 333 1551
> 1.90 1.80 77 206 342 481 701 1033 1403 151 1554
> 1.80 1.72 115 299 490 652 882 1171 1482 61 1543
> 1.72 1.66 190 429 665 844 1054 1347 1555 20 1575
> 1.66 1.60 188 501 762 937 1129 1376 1502 5 1507
> All hkl 638 1673 2727 3648 4897 6962 10022 5539 15561
>
> Shell I/Sigma in resolution shells:
> Lower Upper % of reflections with I / Sigma less than
> limit limit 0 1 2 3 5 10 20 >20 total
> 100.00 3.45 0.1 0.2 0.5 0.8 1.4 3.0 8.0 90.5 98.6
> 3.45 2.74 0.2 0.6 1.5 2.1 3.7 6.8 19.7 79.7 99.4
> 2.74 2.39 0.4 1.2 2.4 3.8 7.1 14.3 37.7 62.0 99.7
> 2.39 2.17 0.5 1.9 4.3 7.7 11.8 24.1 53.4 46.4 99.9
> 2.17 2.02 0.8 3.6 7.6 12.5 18.8 34.2 63.9 35.9 99.8
> 2.02 1.90 2.4 7.7 13.8 20.1 29.6 47.8 78.3 21.4 99.7
> 1.90 1.80 4.9 13.2 22.0 30.9 45.1 66.4 90.2 9.7 99.9
> 1.80 1.72 7.4 19.3 31.7 42.2 57.1 75.7 95.9 3.9 99.8
> 1.72 1.66 12.1 27.2 42.2 53.6 66.9 85.5 98.7 1.3 100.0
> 1.66 1.60 12.2 32.6 49.6 61.0 73.5 89.5 97.7 0.3 98.0
> All hkl 4.1 10.7 17.4 23.3 31.3 44.5 64.1 35.4 99.5
>
> Summary of reflections intensities and R-factors by shells
> R linear = SUM ( ABS(I - <I>)) / SUM (I)
> R square = SUM ( (I - <I>) ** 2) / SUM (I ** 2)
> Chi**2 = SUM ( (I - <I>) ** 2) / (Error ** 2 * N / (N-1) ) )
> In all sums single measurements are excluded
>
> Shell Lower Upper Average Average Norm. Linear Square
> limit Angstrom I error stat. Chi**2 R-fac R-fac
> 100.00 3.45 32741.9 1156.6 362.1 3.738 0.086 0.113
> 3.45 2.74 14355.0 504.8 177.1 3.561 0.090 0.115
> 2.74 2.39 5703.3 232.4 123.1 3.412 0.104 0.121
> 2.39 2.17 4151.1 192.5 120.5 3.053 0.120 0.140
> 2.17 2.02 3216.4 172.0 121.7 2.722 0.129 0.141
> 2.02 1.90 2003.5 141.6 115.5 2.439 0.160 0.168
> 1.90 1.80 1043.6 115.9 105.8 1.771 0.207 0.198
> 1.80 1.72 704.9 113.2 107.4 1.572 0.243 0.230
> 1.72 1.66 503.8 116.9 113.2 1.238 0.280 0.246
> 1.66 1.60 387.1 125.2 122.6 1.092 0.317 0.281
> All reflections 6564.0 289.7 147.5 2.537 0.102 0.114
>
> Intensities of systematic absences
> h k l Intensity Sigma I/Sigma
>
> 0 1 0 0.6 1.1 0.6
> 0 3 0 28.4 18.2 1.6
> 0 5 0 33.9 27.9 1.2
> 0 7 0 -19.3 39.8 -0.5
> 0 9 0 72.3 34.6 2.1
> 0 11 0 57.6 43.1 1.3
> 0 13 0 166.6 48.5 3.4
> 0 15 0 143.7 65.1 2.2
> 0 17 0 101.4 59.2 1.7
> 0 19 0 -87.8 55.2 -1.6
> 0 21 0 -31.5 61.9 -0.5
> 0 23 0 -90.3 74.0 -1.2
> 0 25 0 20.6 94.1 0.2
> 0 27 0 98.8 92.8 1.1
> 0 29 0 -8.8 92.6 -0.1
> 0 31 0 27.2 87.8 0.3
> 0 33 0 -68.5 89.1 -0.8
> -----------
> output for esf of 1.3 and ee of .14 for all shells:
>
> Shell Lower Upper Average Average Norm. Linear Square
> limit Angstrom I error stat. Chi**2 R-fac R-fac
> 100.00 3.45 33071.8 2430.3 335.6 1.855 0.134 0.169
> 3.45 2.74 14530.9 1033.7 164.5 1.833 0.137 0.171
> 2.74 2.39 5790.0 435.7 116.5 1.786 0.144 0.174
> 2.39 2.17 4204.6 333.6 116.0 1.598 0.149 0.176
> 2.17 2.02 3279.2 278.9 118.1 1.495 0.156 0.181
> 2.02 1.90 2045.0 203.5 113.9 1.443 0.178 0.195
> 1.90 1.80 1066.2 144.6 105.5 1.234 0.217 0.215
> 1.80 1.72 721.0 132.2 107.8 1.153 0.248 0.238
> 1.72 1.66 508.1 129.8 114.4 0.957 0.282 0.254
> 1.66 1.60 392.5 135.7 124.5 0.900 0.316 0.280
> All reflections 6679.4 534.1 142.4 1.471 0.143 0.170
>
> --
> --------------------------------------------------------------
> Tim Allison Dept of Biochemistry
> tja2f at dayhoff.med.virginia.edu University of Virginia
You may have very strong MAD signal!
--
Zbyszek Otwinowski | zbyszek at chop.swmed.edu
University of Texas | tel : (214)-648-5098
Southwestern Medical Center | fax : (214)-648-5095
5323 Harry Hines Boulevard Dallas, Texas 75235-9038
[EOB]
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