[BioC] tilingArray getting started questions
noahd at ucla.edu
Tue Sep 7 18:34:22 CEST 2010
I would like to use the tilingArray package to analyze transcript products from several S. cerevisiae mutants using the Affymetrix Tiling Array 1.0R. (Note that these arrays only have one strand of the yeast genome tiled on the array.) I have used these tiling arrays for ChIP-chip experiments. To analyze that data the Starr package was extremely helpful. I have reproduced the examples (using the davidTiling data) in a couple of the tilingArray vignettes and I am getting ready to move into using my data, but have several questions that do not appear in the vignettes or on the archives. The general theme of my questions is if anyone has any experience on the compatibility between Starr and tilingArray.
1. The Starr package has a function (bpmapToProbeAnno) to create a probeAnno object (mapping of the array features to the yeast genome). Can I use this function to create a probeAnno for tilingArray?
2. Reading my CEL files in using Starr (function: readCelFile) can easily create an ExpressionSet object. Should/can I pass this eset to the tilingArray normalizeByReference function after making sure to point to the correct RNA and DNA samples?
3. Alternatively, I could normalize the arrays independently in Starr (function: normalize.Probes (method="loess") and then use the Starr getRatio function to normalize to the "reference" genomic DNA samples. The output of the getRatio function will be an eset that I think I could pass directly to the segChrom function. Am I thinking correctly about this?
4. In David et al, a custom tiling array from Affymetrix was used that had probes corresponding to both strands of DNA. Since I have amplified my single-stranded cDNA with Taq to make a double stranded DNA product which is then hybridized to the array it seems like the tiling arrays that only have one strand (the commercial ones) should work fine for mapping transcripts from both strands. Am I missing something here in the experimental workflow?
If one were to hybridize cDNA directly to an array it follows that both strands of the genome should be represented on the array, but wouldn't lower limit of sensitivity issues then be of concern?
I understand if people have not tried to tie the Starr/Ringo and tilingArray packages together, but I thought I would ask a few questions before spending significant time working out the kinks. Thank you in advance for any help!!
R version 2.11.0 (2010-04-22)
attached base packages:
 grid stats graphics grDevices utils datasets methods base
other attached packages:
 geneplotter_1.26.0 lattice_0.18-5 annotate_1.26.0 RColorBrewer_1.0-2 davidTiling_1.2.12
 GO.db_2.4.1 RSQLite_0.8-4 DBI_0.2-5 AnnotationDbi_1.10.0 tilingArray_1.26.0
 pixmap_0.4-10 Biobase_2.8.0
loaded via a namespace (and not attached):
 affy_1.26.0 affyio_1.16.0 genefilter_1.30.0 limma_3.4.0
 preprocessCore_1.10.0 splines_2.11.0 strucchange_1.4-1 survival_2.35-8
 tools_2.11.0 vsn_3.16.0 xtable_1.5-6
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