[BioC] RE: Advice on print-tip normalization
Gene Cutler
gcutler at amgen.com
Fri Sep 24 22:47:05 CEST 2004
Thanks for everyone's input. I've been looking at the data more
carefully and here's what I've found:
array geometry: grid = 12 rows x 4 cols, block = 28 rows x 27 cols
I see two periodicities.
1) When the intensities are ordered by block column, there are 27
cycles of rising/falling intensities. This gives 1344 spots per cycle
which equals 48 columns of spots per cycle. When these 27 cycles are
overlayed and averaged together, I can see that each 1344 spot cycle
contains another periodicity of 12 cycles: 112 spots per subcycle == 4
columns per subcycle.
2) When the intensities are ordered by block row, there are 12 cycles
of rising/falling intensities. This gives 3024 spots per cycle which
equals 112 rows of spots per cycle or 4 grid blocks per cycle. When
these 12 cycles are overlayed and averaged, I do not see any subcycles.
The coincidence of these cycles with whole row and column numbers
suggests that it is a print tip effect. I don't really understand
print tip geometry yet, so the implications of the numbers above aren't
clear to me. Anyway, the marray boxplot shows what looks like a
similar that goes away after printTipLoess normalization. However, if
I extract the normalized intensities and then plot those, the two
cycles I describe above are still evident.
>>
>> Date: Fri, 17 Sep 2004 10:32:33 -0700
>> From: Gene Cutler <gcutler at amgen.com>
>> Subject: [BioC] Advice on print-tip normalization
>>
>> Hello. I've just started using the marray package for processing a
>> set
>> of spotted oligo arrays. The arrays, when intensities or log ratios
>> are plotted against probe number, show a clear pattern of
>> rising/falling values with a periodicity equal to the grid block size
>> (~3600 spots). I can see a similar periodicity in the printTip
>> boxplots generated with marray. Running printTipLoess smoothes out
>> the
>> boxplot nicely (and the MA plot also looks much nicer), but,
>> surprisingly, when I export the normalized values and plot them
>> against
>> position, the grid block periodicity is little changed.
>>
>> I've tried different span values for the printTipLoess as well as
>> trying with or without scaling (e.g. printTipMAD), but nothing I do
>> seems to have much effect on this data artifact.
>>
>>
Gene Cutler
Research Investigator
Bioinformatics
Amgen SF
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