
Hi Martin

I followed the same steps using the data from a newer version of the illumina pipeline (ver 1.3). Still run into the same issues.

Fuad

##########################
original data illumina 1.3
#########################
> alignQuality(aln)
class: NumericQuality
quality: NA NA ... NA NA (4585850 total)
> head(quality(aln))
class: SFastqQuality
quality:
  A BStringSet instance of length 6
    width seq
[1]    35 ;;;;;;;;;;;;;;;;;;;;;;;;;;:;;;57547
[2]    35 ;:7;;;;;;7;;;;4;;;;;:;:7;77;;;5+444
[3]    35 ;;:;;;:::;;;:;:;;9:6;;;9:;;:;972479
[4]    35 ;;6;;;;;;;;;;:;;;;;;9&9;;;;9;;99587
[5]    35 ;;;;:9;;;;:;;79;;;/::;66;)7;4/78487
[6]    35 ;-;;;;;;;;;;;;;;;;;8;;;;;;;;;055-75
> m <- as(quality(aln), "matrix")
> colMeans(m)
 [1]  -6.044744  -5.936636  -5.924813  -5.958905  -5.956505  -5.921455
 [7]  -5.929669  -5.967875  -5.929279  -5.970600  -5.975925  -6.011533
[13]  -6.462944  -6.520501  -6.601669  -6.651007  -6.805869  -6.832879
[19]  -6.874413  -7.030044  -7.153979  -7.210835  -7.278909  -7.363969
[25]  -7.475453  -7.662229  -7.781669  -7.915972  -8.113778  -8.222790
[31] -12.165080 -12.329542 -12.456710 -12.590711 -12.737429

##########################
incorporated your suggestions
##########################

> qual <- FastqQuality(quality(quality(aln)))
> qual
class: FastqQuality
quality:
  A BStringSet instance of length 4585850
          width seq
      [1]    35 ;;;;;;;;;;;;;;;;;;;;;;;;;;:;;;57547
      [2]    35 ;:7;;;;;;7;;;;4;;;;;:;:7;77;;;5+444
      [3]    35 ;;:;;;:::;;;:;:;;9:6;;;9:;;:;972479
      [4]    35 ;;6;;;;;;;;;;:;;;;;;9&9;;;;9;;99587
      [5]    35 ;;;;:9;;;;:;;79;;;/::;66;)7;4/78487
      [6]    35 ;-;;;;;;;;;;;;;;;;;8;;;;;;;;;055-75
      [7]    35 ;:9;;;;;;;;;;;;;:;:;;;;:;;;;+:54455
      [8]    35 ;;;;;;:;;;:;;;;;:;;3:9;;;;:;;;45544
      [9]    35 ;;:;;;;;;;4;;;4;:7;:;;;74;;;;644015
      ...   ... ...
[4585842]    35 0:;7;;-;;;;;:;;7)-.25;;&;;9;7;.4444
[4585843]    35 7477:.-;9*;3;0979489.-9::&;49:5-344
[4585844]    35 6:;::;;;;;:;:;;6::98:3962:949:24424
[4585845]    35 0:;;:::6;::.:::626:277:8)9::7:)5--0
[4585846]    35 4;7+9-;;;;6:;);9;+2037:&0-270&4.-1.
[4585847]    35 :;;::;;;;;;;0;:96.:9:9;2;);99;54544
[4585848]    35 ;;:9;:;;;;:;:7,0)9;);))9.;6:4;4245-
[4585849]    35 8:;;;;;:668:;2;:,836:;69:993::454,0
[4585850]    35 889;:;;;;;;-;-.89);;;);;4044*7*4.41
> initialize(aln, quality=qual)
class: AlignedRead
length: 4585850 reads; width: 35 cycles
chromosome: shank3genomic shank3genomic ... shank3genomic shank3genomic 
position: 37625 19915 ... 13278 39832 
strand: + + ... + + 
alignQuality: NumericQuality 
alignData varLabels: similar mismatch

> m <- as(quality(aln), "matrix")
> colMeans(m)
 [1]  -6.044744  -5.936636  -5.924813  -5.958905  -5.956505  -5.921455
 [7]  -5.929669  -5.967875  -5.929279  -5.970600  -5.975925  -6.011533
[13]  -6.462944  -6.520501  -6.601669  -6.651007  -6.805869  -6.832879
[19]  -6.874413  -7.030044  -7.153979  -7.210835  -7.278909  -7.363969
[25]  -7.475453  -7.662229  -7.781669  -7.915972  -8.113778  -8.222790
[31] -12.165080 -12.329542 -12.456710 -12.590711 -12.737429

> alignQuality(aln)
class: NumericQuality
quality: NA NA ... NA NA (4585850 total)

###############################
Also tried your second suggestion and ended up with teh following error
##############################

> m <- as(FastqQuality(quality(quality(aln)), "matrix"))
Error in .identC(Class, "double") : 
  element 2 is empty;
   the part of the args list of '.Call' being evaluated was:
   ("R_identC", c1, c2, PACKAGE = "methods")



> sessionInfo()
R version 2.9.1 (2009-06-26) 
x86_64-unknown-linux-gnu 

locale:
LC_CTYPE=en_US.UTF-8;LC_NUMERIC=C;LC_TIME=en_US.UTF-8;LC_COLLATE=en_US.UTF-8;LC_MONETARY=C;LC_MESSAGES=en_US.UTF-8;LC_PAPER=en_US.UTF-8;LC_NAME=C;LC_ADDRESS=C;LC_TELEPHONE=C;LC_MEASUREMENT=en_US.UTF-8;LC_IDENTIFICATION=C

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

other attached packages:
[1] ShortRead_1.3.22   lattice_0.17-25    BSgenome_1.13.10   Biostrings_2.13.29
[5] IRanges_1.3.44    

loaded via a namespace (and not attached):
[1] Biobase_2.4.1 grid_2.9.1    hwriter_1.1  



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