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Question: Problem with reading raw Illumina Human HT-12 v4 data using beadarray
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gravatar for jaro.slamecka
2.7 years ago by
Mitchell Cancer Institute, Mobile AL, USA
jaro.slamecka30 wrote:

Hi,

I am new to microarray data analysis, trying to put the pieces of puzzle together. Trying to read raw Illumina Human HT-12 v4 data using beadarray. My raw data folder contains csv, txt, EIF, idat, locs, tif, xml file for each of the 12 samples. There is one sdf file, one metrics.txt file and 3 fiducial #N_Red_center.tif files.

Function readIllumina(useImages = FALSE, illuminaAnnotation = "Humanv4") gives the error below. Each txt file contains only two columns: Code Grn. Is the format of my raw data unsuitable for readIllumina? Or is there a way to instruct it so that it read the data properly? Thanks in advance!

Trying to read all Illumina files inD:\RData\HA1
Processing section 9992405085_A
Error in `[.data.frame`(lines, 1, 1:4) : undefined columns selected

> traceback()
6: stop("undefined columns selected")
5: `[.data.frame`(lines, 1, 1:4)
4: lines[1, 1:4]
3: numberOfColumns(file, sep = sep)
2: readBeadLevelTextFile(file.path(targets$directory[i], targets$textFile[i]),
       dec = dec)
1: readIllumina(useImages = FALSE, illuminaAnnotation = "Humanv4")

> sessionInfo()
R version 3.1.2 (2014-10-31)
Platform: x86_64-w64-mingw32/x64 (64-bit)

locale:
[1] LC_COLLATE=English_United States.1252
[2] LC_CTYPE=English_United States.1252   
[3] LC_MONETARY=English_United States.1252
[4] LC_NUMERIC=C                          
[5] LC_TIME=English_United States.1252    

attached base packages:
[1] parallel  stats     graphics  grDevices utils     datasets  methods  
[8] base     

other attached packages:
[1] beadarray_2.16.0    ggplot2_1.0.0       GEOquery_2.32.0    
[4] lumi_2.18.0         Biobase_2.26.0      BiocGenerics_0.12.1

loaded via a namespace (and not attached):
 [1] affy_1.44.0             affyio_1.34.0           annotate_1.44.0        
 [4] AnnotationDbi_1.28.1    base64_1.1              base64enc_0.1-2        
 [7] BatchJobs_1.5           BBmisc_1.8              BeadDataPackR_1.18.0   
[10] beanplot_1.2            BiocInstaller_1.16.1    BiocParallel_1.0.0     
[13] biomaRt_2.22.0          Biostrings_2.34.1       bitops_1.0-6           
[16] brew_1.0-6              bumphunter_1.6.0        checkmate_1.5.1        
[19] codetools_0.2-10        colorspace_1.2-4        DBI_0.3.1              
[22] digest_0.6.8            doRNG_1.6               fail_1.2               
[25] foreach_1.4.2           genefilter_1.48.1       GenomeInfoDb_1.2.4     
[28] GenomicAlignments_1.2.1 GenomicFeatures_1.18.3  GenomicRanges_1.18.4   
[31] grid_3.1.2              gtable_0.1.2            illuminaio_0.8.0       
[34] IRanges_2.0.1           iterators_1.0.7         KernSmooth_2.23-13     
[37] lattice_0.20-29         limma_3.22.4            locfit_1.5-9.1         
[40] MASS_7.3-37             Matrix_1.1-5            matrixStats_0.13.1     
[43] mclust_4.4              methylumi_2.12.0        mgcv_1.8-4             
[46] minfi_1.12.0            multtest_2.22.0         munsell_0.4.2          
[49] nleqslv_2.5             nlme_3.1-119            nor1mix_1.2-0          
[52] pkgmaker_0.22           plyr_1.8.1              preprocessCore_1.28.0  
[55] proto_0.3-10            quadprog_1.5-5          R.methodsS3_1.6.1      
[58] RColorBrewer_1.1-2      Rcpp_0.11.4             RCurl_1.95-4.5         
[61] registry_0.2            reshape_0.8.5           reshape2_1.4.1         
[64] rngtools_1.2.4          Rsamtools_1.18.2        RSQLite_1.0.0          
[67] rtracklayer_1.26.2      S4Vectors_0.4.0         scales_0.2.4           
[70] sendmailR_1.2-1         siggenes_1.40.0         splines_3.1.2          
[73] stats4_3.1.2            stringr_0.6.2           survival_2.37-7        
[76] tools_3.1.2             XML_3.98-1.1            xtable_1.7-4           
[79] XVector_0.6.0           zlibbioc_1.12.0

 

Jaro

Jaroslav Slamecka, PhD.
Mitchell Cancer Institute
University of South Alabama
1660 Springhill Ave.
Mobile, AL 36604

ADD COMMENTlink modified 2.7 years ago • written 2.7 years ago by jaro.slamecka30
0
gravatar for Mike Smith
2.7 years ago by
Mike Smith2.1k
EMBL Heidelberg / de.NBI
Mike Smith2.1k wrote:

Hi Jaro,

Thanks for bringing this to our attention.  Unfortunately, if your txt file only has two columns you can't use many of the features of beadarray (e.g. BASH) that rely on the coordinate information, and the software currently fails to read the data (admittedly with a not very helpful message).

That said there are still some advantages to working with bead-level data over Illumina's summarized version, such as being able to perform outlier removal on the log2 scale.  For the moment, if you wish to use the beadarray package you could fake the two coordinate columns by giving everything a value of 0.  beadarray should be able to read that, but you have to be careful not to make use of any of the spatial information as this would obviously be misleading.

If you have access to the iScan machine and want to ensure that the X,Y coordinates are included in the future, the instructions can be found on page 35 of the user guide here:

http://support.illumina.com/content/dam/illumina-support/documents/myillumina/f1805681-577a-4db3-9240-333ac009d333/iscan_system_user_guide_11313539_b.pdf

Let me know if you need anything more,

Mike

ADD COMMENTlink written 2.7 years ago by Mike Smith2.1k
0
gravatar for Gordon Smyth
2.7 years ago by
Gordon Smyth32k
Walter and Eliza Hall Institute of Medical Research, Melbourne, Australia
Gordon Smyth32k wrote:

You could try read.idat() in the limma package, which will read in the expression data from the idat files.

ADD COMMENTlink written 2.7 years ago by Gordon Smyth32k
0
gravatar for jaro.slamecka
2.7 years ago by
Mitchell Cancer Institute, Mobile AL, USA
jaro.slamecka30 wrote:

Thank you both so much for your help! Too bad the core lab didn't set the iScan to include the XY coordinate data, I'll make sure they do the next time. And I will look into read.idat in limma.

Jaro

ADD COMMENTlink written 2.7 years ago by jaro.slamecka30
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