Dear all,

this is a follow up from my last question. When my run my DESeq2 analysis using the usual estimateSizeFactors, it runs fine, however if I provide a normalisation matrix, I get the following error on estimateDispersions:

Error in fitBetaWrapper(ySEXP = counts(object), xSEXP = modelMatrix, nfSEXP = normalizationFactors,  : in call to fitBeta, the following arguments contain NA: alpha_hatSEXP

I have generated the matrix & checked for the absence of NA and 0 & run DESeq2 as follows:

data <- read.table(file = "PATH.tsv", header=TRUE, row.name=1)
normFactors <- data.matrix(data)
normFactors <- normFactors / exp(rowMeans(log(normFactors)))
normFactors[is.na(normFactors)] <- 1
dds = DESeqDataSetFromMatrix(experiment, design, design = ~ genotype)
normalizationFactors(dds) <- normFactors
dds <- estimateDispersions(dds)

My design file shows that there are >3 replicates per group

   track group    batch genotype
1  A.12.R1 A-12   2nd       A
2  A.12.R2 A-12   2nd       A
3  A.12.R3 A-12   2nd       A
4  A.12.R4 A-12   2nd       A
5  B.12.R1 B-12   2nd       B
6  B.12.R2 B-12   2nd       B
7  B.12.R3 B-12   2nd       B 
8  B.12.R4 C-12   2nd       C  
9  C.12.R1 C-12   2nd       C  
10 C.12.R2 A-12   2nd       A 
11 C.12.R3 A-12   2nd       A  
12 C.12.R4 C-12   2nd       C   

Many thanks, Jakub

session.Info():

R version 3.2.3 (2015-12-10)
Platform: x86_64-pc-linux-gnu (64-bit)
Running under: Scientific Linux release 6.7 (Carbon)

locale:
[1] C

attached base packages:
 [1] grid      parallel  stats4    tools     stats     graphics  grDevices
 [8] utils     datasets  methods   base     

other attached packages:
 [1] biomaRt_2.26.1                          
 [2] geneplotter_1.48.0                      
 [3] annotate_1.48.0                         
 [4] XML_3.98-1.4                            
 [5] lattice_0.20-33                         
 [6] MatchIt_2.4-21                          
 [7] MASS_7.3-45                             
 [8] TxDb.Mmusculus.UCSC.mm10.knownGene_3.2.2
 [9] TxDb.Mmusculus.UCSC.mm10.ensGene_3.2.2  
[10] GenomicFeatures_1.22.13                 
[11] AnnotationDbi_1.32.3                    
[12] VennDiagram_1.6.16                      
[13] futile.logger_1.4.1                     
[14] goseq_1.22.0                            
[15] RSQLite_1.0.0                           
[16] DBI_0.3.1                               
[17] geneLenDataBase_1.6.0                   
[18] BiasedUrn_1.07                          
[19] vsn_3.38.0                              
[20] dynamicTreeCut_1.63-1                   
[21] ggplot2_2.1.0                           
[22] gplots_3.0.1                            
[23] RColorBrewer_1.1-2                      
[24] DESeq2_1.10.1                           
[25] RcppArmadillo_0.6.600.4.0               
[26] Rcpp_0.12.4                             
[27] SummarizedExperiment_1.0.2              
[28] Biobase_2.30.0                          
[29] GenomicRanges_1.22.4                    
[30] GenomeInfoDb_1.6.3                      
[31] IRanges_2.4.8                           
[32] S4Vectors_0.8.11                        
[33] BiocGenerics_0.16.1                     

loaded via a namespace (and not attached):
 [1] locfit_1.5-9.1          GO.db_3.2.2             Rsamtools_1.22.0       
 [4] Biostrings_2.38.4       gtools_3.5.0            plyr_1.8.3             
 [7] futile.options_1.0.0    acepack_1.3-3.3         BiocInstaller_1.20.1   
[10] zlibbioc_1.16.0         gdata_2.17.0            Matrix_1.2-4           
[13] rpart_4.1-10            preprocessCore_1.32.0   splines_3.2.3          
[16] BiocParallel_1.4.3      foreign_0.8-66          RCurl_1.95-4.8         
[19] munsell_0.4.3           rtracklayer_1.30.2      mgcv_1.8-12            
[22] nnet_7.3-12             gridExtra_2.2.1         Hmisc_3.17-3           
[25] GenomicAlignments_1.6.3 bitops_1.0-6            nlme_3.1-126           
[28] xtable_1.8-2            gtable_0.2.0            affy_1.48.0            
[31] scales_0.4.0            KernSmooth_2.23-15      XVector_0.10.0         
[34] genefilter_1.52.1       affyio_1.40.0           limma_3.26.8           
[37] latticeExtra_0.6-28     Formula_1.2-1           lambda.r_1.1.7         
[40] survival_2.38-3         colorspace_1.2-6        cluster_2.0.3          
[43] caTools_1.17.1