DEXSeq Error loading in data set - subscript out of bound
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Entering edit mode
6 days ago
osiemen ▴ 20

Hi I am currently following the tutorial of DEXSeq to do a differential Exon usage analysis on my dataset. I am currently at the step where I read my dataset in to R to construct a DEXSeqDataSet using the function DEXSeqDataSetFromHTSeq() .

dxd = DEXSeqDataSetFromHTSeq(
   countFiles,
   sampleData=sampleTable,
   design= ~ sample + exon + condition:exon,
   flattenedfile=flattenedFile )

I have total 4 countFiles which I constructed from my actual dataset to apply this function on a smaller dataset. The countFiles look like following:

RNA1_test.txt:

""ENSG00000000003.14"":"001"    36
""ENSG00000000003.14"":"002"    183
""ENSG00000000003.14"":"003"    45
""ENSG00000000003.14"":"004"    0
""ENSG00000000003.14"":"005"    32
""ENSG00000000003.14"":"006"    21
""ENSG00000000003.14"":"007"    22
""ENSG00000000003.14"":"008"    24
""ENSG00000000003.14"":"009"    26
""ENSG00000000003.14"":"010"    17
""ENSG00000000003.14"":"011"    34
""ENSG00000000003.14"":"012"    10
""ENSG00000000003.14"":"013"    12
""ENSG00000000003.14"":"014"    7
""ENSG00000000003.14"":"015"    2
""ENSG00000000003.14"":"016"    0
""ENSG00000000003.14"":"017"    0

RNA2_test.txt:

""ENSG00000000003.14"":"001"    34
""ENSG00000000003.14"":"002"    191
""ENSG00000000003.14"":"003"    60
""ENSG00000000003.14"":"004"    0
""ENSG00000000003.14"":"005"    41
""ENSG00000000003.14"":"006"    36
""ENSG00000000003.14"":"007"    42
""ENSG00000000003.14"":"008"    50
""ENSG00000000003.14"":"009"    47
""ENSG00000000003.14"":"010"    26
""ENSG00000000003.14"":"011"    38
""ENSG00000000003.14"":"012"    5
""ENSG00000000003.14"":"013"    18
""ENSG00000000003.14"":"014"    7
""ENSG00000000003.14"":"015"    1
""ENSG00000000003.14"":"016"    1
""ENSG00000000003.14"":"017"    0

etc...

The amount of lines the transcripts/Exons in the the 4 countFiles are identical only the counts are different. I also used a sample of the actual flattened gff file that was used for the counting the reads :

chrX    dexseq_prepare_annotation.py    aggregate_gene  100627109       100639991       .       -       .       gene_id ""ENSG00000000003.14""
chrX    dexseq_prepare_annotation.py    exonic_part     100627109       100628669       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000612152.4""; exonic_part_number "001"
chrX    dexseq_prepare_annotation.py    exonic_part     100628670       100629986       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000373020.8"+"ENST00000612152.4""; exonic_part_number "002"
chrX    dexseq_prepare_annotation.py    exonic_part     100630759       100630866       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000373020.8"+"ENST00000612152.4""; exonic_part_number "003"
chrX    dexseq_prepare_annotation.py    exonic_part     100632063       100632068       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000614008.4""; exonic_part_number "004"
chrX    dexseq_prepare_annotation.py    exonic_part     100632485       100632540       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000373020.8"+"ENST00000614008.4""; exonic_part_number "005"
chrX    dexseq_prepare_annotation.py    exonic_part     100632541       100632568       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000373020.8"+"ENST00000496771.5"+"ENST00000614008.4""; exonic_part_number "006"
chrX    dexseq_prepare_annotation.py    exonic_part     100633405       100633441       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000373020.8"+"ENST00000496771.5"+"ENST00000614008.4"+"ENST00000612152.4""; exonic_part_number "007"
chrX    dexseq_prepare_annotation.py    exonic_part     100633442       100633539       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000614008.4"+"ENST00000612152.4"+"ENST00000494424.1"+"ENST00000496771.5"+"ENST00000373020.8""; exonic_part_number "008"
chrX    dexseq_prepare_annotation.py    exonic_part     100633931       100634029       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000614008.4"+"ENST00000612152.4"+"ENST00000494424.1"+"ENST00000496771.5"+"ENST00000373020.8""; exonic_part_number "009"
chrX    dexseq_prepare_annotation.py    exonic_part     100635178       100635252       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000614008.4"+"ENST00000612152.4"+"ENST00000494424.1"+"ENST00000496771.5"+"ENST00000373020.8""; exonic_part_number "010"
chrX    dexseq_prepare_annotation.py    exonic_part     100635558       100635746       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000614008.4"+"ENST00000612152.4"+"ENST00000494424.1"+"ENST00000496771.5"+"ENST00000373020.8""; exonic_part_number "011"
chrX    dexseq_prepare_annotation.py    exonic_part     100636191       100636607       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000496771.5""; exonic_part_number "012"
chrX    dexseq_prepare_annotation.py    exonic_part     100636608       100636689       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000373020.8"+"ENST00000496771.5""; exonic_part_number "013"
chrX    dexseq_prepare_annotation.py    exonic_part     100636690       100636792       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000373020.8""; exonic_part_number "014"
chrX    dexseq_prepare_annotation.py    exonic_part     100636793       100636806       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000373020.8"+"ENST00000614008.4"+"ENST00000494424.1"+"ENST00000612152.4""; exonic_part_number "015"
chrX    dexseq_prepare_annotation.py    exonic_part     100636807       100637104       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000614008.4"+"ENST00000494424.1"+"ENST00000612152.4""; exonic_part_number "016"
chrX    dexseq_prepare_annotation.py    exonic_part     100639945       100639991       .       -       .       gene_id ""ENSG00000000003.14""; transcripts ""ENST00000494424.1""; exonic_part_number "017"

However when I run my code :

#get count file names
countFiles= list.files(path = "Desktop",pattern = "RNA", full.names = TRUE)
basename(countFiles)

[1] "RNA1_test.txt" "RNA2_test.txt" "RNA3_test.txt" "RNA4_test.txt"
#get gff file name
 flattenedFile = list.files(path = "Desktop", pattern="gff$", full.names=TRUE)
basename(flattenedFile)
[1] "dexseq_test.gff"

sampleTable = data.frame(
  row.names = c( "retina1", "retina2", 
                 "brain1", "brain2"  ),
  condition = c("experiment", "experiment",
                "control", "control" ))

dxd = DEXSeqDataSetFromHTSeq(
countFiles,
sampleData=sampleTable2,
design= ~ sample + exon + condition:exon,
flattenedfile=flattenedFile )

I receive the following error:

Error in FUN(X[[i]], ...) : subscript out of bound

I am not sure what might cause this error, since the gene ids in the gff file also match the ones used in my countFiles. Anyone familiar with this error?

DEXSeq Exon RNA-Seq Expression Count • 120 views
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Entering edit mode

One of the differences I noticed between my files and the files used in the example are the quotes " " in the countFiles -> ""ENSG00000000003.14"":"001" 34 , but im not sure if that could cause it

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