I am working with a custom chip which does not have mm probes, but which does have negative control probes. I would like to see the results of gcrma adjustment of the probe values. I have thus chosen all of the indices of the negative controls and am trying to use them as described in the gcrma package: The indices are here: > randomprobeindexes[1:5,] x y [1,] 114 117 [2,] 116 117 [3,] 118 117 [4,] 120 117 [5,] 122 117 > I now use them to do gcrma: > bg.adjust.gcrma(newdata, affinity.source="local", type="affinities", NCprobe=randomprobeindexes) Adjusting for optical effect.Error in exprs(abatch)[Index, i] <- exprs(abatch)[Index, i] - min(exprs(abatch)[Index, : NAs are not allowed in subscripted assignments > This is, I think, due to this: > mm(newdata)[200:205,] dmso5.CEL mnng5.CEL mock5.CEL uv5.CEL [1,] NA NA NA NA [2,] NA NA NA NA [3,] NA NA NA NA [4,] NA NA NA NA [5,] NA NA NA NA [6,] NA NA NA NA > And: > mmindex(newdata)[200:202] $`AFFX-yel006_5_copy5_at` [1] NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA $`AFFX-yel006_5_copy6_at` [1] NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA $`AFFX-yel006_M_copy1_at` [1] NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA > We do have some mm values on the chip, since there is always one control set present which does have mm probes with it. This one is required for affymetrix own purposes. Besides this there are no mm values. which means that most of the mmindex list just states na, and as shown above, so does the mm list too. Now, my initial strategy was to try and replace all mm values with zero. gcrma does not use mm values when negative control indices are given, so this should, afaik, be ok. However, I am not able to do this, and I think this is due to me not having any mm indices to use for the replacement. Does any of you have any tips on how to overcome this? TIA, Karin -- Karin Lagesen, PhD student karin.lagesen at medisin.uio.no http://folk.uio.no/karinlag

Hi Karin, You might look at a similar thread from a year ago: http://article.gmane.org/gmane.science.biology.informatics.conductor/3 997/match=gcrma+mm+probe I think the take home message was that you can either use bg.parameters.ns() to estimate the background, or you can assume that most genes are not expressed, in which case you can pass the PM as MM values. Either way will require some work on your part to figure out how to get things to work. Best, Jim Karin Lagesen wrote: > > I am working with a custom chip which does not have mm probes, > but which does have negative control probes. I would like to > see the results of gcrma adjustment of the probe values. > > I have thus chosen all of the indices of the negative controls > and am trying to use them as described in the gcrma package: > > The indices are here: > > > >>randomprobeindexes[1:5,] > > x y > [1,] 114 117 > [2,] 116 117 > [3,] 118 117 > [4,] 120 117 > [5,] 122 117 > > > I now use them to do gcrma: > > >>bg.adjust.gcrma(newdata, affinity.source="local", type="affinities", NCprobe=randomprobeindexes) > > Adjusting for optical effect.Error in exprs(abatch)[Index, i] <- exprs(abatch)[Index, i] - min(exprs(abatch)[Index, : > NAs are not allowed in subscripted assignments > > > This is, I think, due to this: > > >>mm(newdata)[200:205,] > > dmso5.CEL mnng5.CEL mock5.CEL uv5.CEL > [1,] NA NA NA NA > [2,] NA NA NA NA > [3,] NA NA NA NA > [4,] NA NA NA NA > [5,] NA NA NA NA > [6,] NA NA NA NA > > > And: > > >>mmindex(newdata)[200:202] > > $`AFFX-yel006_5_copy5_at` > [1] NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA > > $`AFFX-yel006_5_copy6_at` > [1] NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA > > $`AFFX-yel006_M_copy1_at` > [1] NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA NA > > > > We do have some mm values on the chip, since there is always one > control set present which does have mm probes with it. This one > is required for affymetrix own purposes. Besides this there are > no mm values. which means that most of the mmindex list just > states na, and as shown above, so does the mm list too. > > Now, my initial strategy was to try and replace all mm values > with zero. gcrma does not use mm values when negative control > indices are given, so this should, afaik, be ok. However, I > am not able to do this, and I think this is due to me not > having any mm indices to use for the replacement. > > Does any of you have any tips on how to overcome this? > > TIA, > > Karin -- James W. MacDonald, M.S. Biostatistician Affymetrix and cDNA Microarray Core University of Michigan Cancer Center 1500 E. Medical Center Drive 7410 CCGC Ann Arbor MI 48109 734-647-5623 ********************************************************** Electronic Mail is not secure, may not be read every day, and should not be used for urgent or sensitive issues.