[Bioc-sig-seq] Problem with ShortRead reading quality scores from bowtie

[Martin Morgan]

Martin Morgan wrote:
> Fuad Gwadry wrote:
>> Hi All
>>
>>  
>>
>> I am getting negative values when reading quality scores when I read 
>> data generated in bowtie. Has anyone run into the same issue when 
>> using data generated by bowtie ? My session info is below. 
> 
> Hi Fuad -- ShortRead is reading the quality scores on the wrong scale 
> (solexa, rather than phred; this will be fixed before the next release). 
> Try
> 
>   qual <- FastqQuality(quality(quality(aln))
>   initialize(aln, quality=qual)
> 
> to update aln, or
> 
>   m <- as(FastqQuality(quality(quality(aln)), "matrix")
> 
> for a one-off solution.

I wanted to clarify, too, both for this post and one yesterday, that 
as() is simply converting the character encoding to the corresponding 
integer value that each letter encodes; there is a secondary mapping 
from this encoding to log-odds or phred score that is not being 
performed. This step is, I think

   10^(-m/10) for phred scores
   1 - 1 / (1 + 10^(-m/10)) for Solexa scores

Solexa has changed its encoding scheme very recently; I think it is now 
standard phred but am not sure.

Martin

> 
> Martin
> 
>>
>>  
>>
>> Thanks in advance
>>
>>  
>>
>> Fuad
>>
>>  
>>
>>> aln
>> class: AlignedRead
>> length: 4591807 reads; width: 32 cycles
>> chromosome: chr13 chr7 ... chr6 chr4 position: 93437004 13223395 ... 
>> 23636747 23353864 strand: - - ... + + alignQuality: NumericQuality 
>> alignData varLabels: similar mismatch
>>
>>> m <- as(quality(aln), "matrix")
>>> colMeans(m)
>>  [1]  -7.186638  -7.205858  -7.203382  -7.197175  -7.203629  -7.217016
>>  [7]  -7.240661  -7.249238  -7.268499  -7.286551  -7.306615  -7.324003
>> [13]  -7.523238  -7.581242  -7.697591  -7.695861  -7.735321  -7.743323
>> [19]  -7.752996  -7.849403  -7.862658  -7.931969  -7.979778  -8.029288
>> [25]  -8.120469  -8.215818  -8.335176  -8.411609  -8.587005  -8.820979
>> [31] -11.447326 -11.644198
>>
>>
>>> 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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> 
> 


-- 
Martin Morgan
Computational Biology / Fred Hutchinson Cancer Research Center
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PO Box 19024 Seattle, WA 98109

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