Hi, I am doing some Illumina analysis using HumanWG-6_V2 microarrays, and have been using the annotation file: HumanWG-6_V2_0_R4_11223189_A.bgx, and I am normalising using the NEQC function in the LIMMA package. I know there are traditionally a number of Illumina identifiers and I am concerned that I may have potentially been using the wrong ones, and I'm not sure whether this has affected the normalisation proceedure, or anything at all. After summarisation in Genome Studio, when looking at the 'Sample Probe Profile', the main identifiers that come up (and which I have used in LIMMA) are 'PROBE_ID' and 'SYMBOL', the first row being ILMN_1762337 and 7A5 respectively. I also noticed that this PROBE_ID column was the one used in the Illumina example in the LIMMA manual. HOWEVER, in Genome Studio, there is also a column called 'ProbeID'. This does not exist in the original annotation file (HumanWG-6_V2_0_R4_11223189_A), but it is identical to the Array_Address_ID (except for the preceeding 000s), the latter of which is both in Genome Strudio and in the Annotation file, and UNIQUE to the version of the microarray. IN CONTRAST, in the 'Control Probe Profile' in Genome Studio, there is only the 'TargetID' and the 'ProbeID' available, the latter of which (I believe) is the Array_Address_ID? HENCE, for the LIMMA input, I am wondering whether I am correct when I have included the Sample Probe ID text file (which includes PROBE_ID, that is, ILMN_1762337), and the Control Probe ID text file (which includes ProbeID instead, which is most likely the Array Address ID). Many thanks in advance, William d'Avigdor

Dear William, What you have done is correct. As you have found, the 'ProbeID' is the same as the Array_Address_ID. The 'ProbeID' column was used in the old versions of Illumina BeadChip arrays, and it was later replaced with 'PROBE_ID" in the newer versions of BeadChips. The neqc() function uses negative control probes to carry out background correction. The 'TargetID' column in the control probe profile file indicates the types of control probes and the negative control probes have the type of 'NEGATIVE'. Neqc also uses all the probes including regular probes and all types of control probes (negative controls, housekeeping, ...) to perform a quantile between- array normalization. Best wishes, Wei On Jul 2, 2013, at 3:56 PM, William D'Avigdor wrote: > Hi, > > I am doing some Illumina analysis using HumanWG-6_V2 microarrays, and have been using the annotation file: HumanWG- 6_V2_0_R4_11223189_A.bgx, and I am normalising using the NEQC function in the LIMMA package. > > I know there are traditionally a number of Illumina identifiers and I am concerned that I may have potentially been using the wrong ones, and I'm not sure whether this has affected the normalisation proceedure, or anything at all. > > After summarisation in Genome Studio, when looking at the 'Sample Probe Profile', the main identifiers that come up (and which I have used in LIMMA) are 'PROBE_ID' and 'SYMBOL', the first row being ILMN_1762337 and 7A5 respectively. I also noticed that this PROBE_ID column was the one used in the Illumina example in the LIMMA manual. > > HOWEVER, in Genome Studio, there is also a column called 'ProbeID'. This does not exist in the original annotation file (HumanWG- 6_V2_0_R4_11223189_A), but it is identical to the Array_Address_ID (except for the preceeding 000s), the latter of which is both in Genome Strudio and in the Annotation file, and UNIQUE to the version of the microarray. > > IN CONTRAST, in the 'Control Probe Profile' in Genome Studio, there is only the 'TargetID' and the 'ProbeID' available, the latter of which (I believe) is the Array_Address_ID? > > HENCE, for the LIMMA input, I am wondering whether I am correct when I have included the Sample Probe ID text file (which includes PROBE_ID, that is, ILMN_1762337), and the Control Probe ID text file (which includes ProbeID instead, which is most likely the Array Address ID). > > Many thanks in advance, > William d'Avigdor > > _______________________________________________ > Bioconductor mailing list > Bioconductor at r-project.org > https://stat.ethz.ch/mailman/listinfo/bioconductor > Search the archives: http://news.gmane.org/gmane.science.biology.informatics.conductor ______________________________________________________________________ The information in this email is confidential and intend...{{dropped:6}}