[BioC] Bizarre artifacts in MA plots for ChIP-chip data

[Steve Lianoglou]

Hi all,

As the subject suggests, I'm getting some weird artifacts in ChIP-chip  
data I'm looking at (Aiglent 244k human promoter). This is my first  
foray into ChIP-chip analysis, so I'm sorry if these are naive  
questions.

Instead of painting a picture w/ a 1000 words, I figure it would be  
best to drop a few pictures online if someone has some time to look at  
them and can offer some suggestion as to what might be happening.

I have 3 replicates experiments for two different Transcription  
Factors. In both cases, two of the three replicates look real screwy,  
and 1 of them (for both TFs) looks somehow "ok". The plots shown below  
are using the r/gProcessedSignals that from Agilent's feature  
extraction software, so their error-corrected/bg-subtracted is being  
used. I've also removed the control probes.

For TF1:
The "faux" microarray image looks fine. Here's pic of the Log2 ratios  
between the red/green channels:
http://cbio.mskcc.org/~lianos/files/bioconductor/bad1.Log2Ratio.png

Here's the funky MA plot for the same replicate expt:
http://cbio.mskcc.org/~lianos/files/bioconductor/bad1.png

Faux microarray image for "better" replicate of same TF:
http://cbio.mskcc.org/~lianos/files/bioconductor/better1.Log2Ratio.png

MA plot for the better replicate of TF1:
http://cbio.mskcc.org/~lianos/files/bioconductor/better1.png

You can see that at low the low-A values of the plot, there is a  
pretty significant density of probes that skew the M values quite  
high. They are also there in a smaller amount on the rep2 picture,  
(but below the M=0 line).

Does anyone have an intuition as to what might be causing this? I  
don't have die-swap experiments, so I'm not sure if it's dye bias.  
Even so, is there such thing as intensity-specific dye-bias (to  
explain it only happening at low values of A (roughly A < 7)?

The effect is even more exacerbated in TF2:
Here's a "weird" replicate:
http://cbio.mskcc.org/~lianos/files/bioconductor/bad2.png

Here's a more reasonable replicate for TF2:
http://cbio.mskcc.org/~lianos/files/bioconductor/better2.png

The faux log2-ratio images for these arrays look much like the other  
ones, ie. there are no obvious smudges, etc.

I'm going to check out what probes correspond to those dark spots in  
the low-A range, but in the meantime I was wondering if anybody had  
any suggestions.

Thanks for any help,
-steve

--
Steve Lianoglou
Graduate Student: Physiology, Biophysics and Systems Biology
Weill Cornell Medical College of Cornell University

http://cbio.mskcc.org/~lianos