Hi, I have got an Error in .local(.Object, ...) : allGenes must be a factor with 2 levels using the following code
> library(edgeR)
> library(topGO)
...
> tr <- glmTreat(fit, contrast=B.LvsP, lfc=log2(1.5))
> topTags(tr)
Coefficient: -1*Leaves.2 1*Leaves.3
logFC unshrunk.logFC logCPM PValue FDR
sp0090975 2.6 2.6 2.8 5.7e-14 2.7e-09
sp0037632 -3.0 -3.0 3.4 1.7e-13 2.7e-09
sp0074153 -3.9 -3.9 3.8 1.8e-13 2.7e-09
sp0008306 3.2 3.2 2.9 1.8e-13 2.7e-09
sp0073530 -4.5 -4.5 3.4 2.3e-12 2.3e-08
sp0025713 -3.9 -3.9 4.4 2.6e-12 2.3e-08
sp0037721 7.8 8.0 2.4 2.7e-12 2.3e-08
sp0083660 2.0 2.0 4.4 3.2e-12 2.3e-08
sp0052245 -2.9 -2.9 4.9 3.9e-12 2.6e-08
sp0071520 -3.3 -3.3 2.7 5.8e-12 3.4e-08
> geneID2GO <- readMappings(file = "~GOmapping.tsv")
> head(geneID2GO)
$`"V1"`
[1] "\"V14\""
$`"sp0000005"`
[1] "\"GO:0003723\""
$`"sp0000006"`
[1] "\"GO:0016021\""
$`"sp0000007"`
[1] "\"GO:0003700" "GO:0006355" "GO:0043565\""
$`"sp0000016"`
[1] "\"GO:0046983\""
$`"sp0000017"`
[1] "\"GO:0004672" "GO:0005524" "GO:0006468\""
> geneUniverse <- names(geneID2GO)
> head(geneUniverse)
[1] "\"V1\"" "\"sp0000005\"" "\"sp0000006\"" "\"sp0000007\"" "\"sp0000016\""
[6] "\"sp0000017\""
> genesOfInterest <- rownames(tr@.Data[[1]])
> head(genesOfInterest)
[1] "sp0025247" "sp0025250" "sp0025268" "sp0025270" "sp0025282" "sp0056834"
> geneList <- factor(as.integer(geneUniverse %in% genesOfInterest))
> head(geneList)
[1] 0 0 0 0 0 0
Levels: 0
> names(geneList) <- geneUniverse
> myGOdata <- new("topGOdata",
+ description="My project",
+ ontology="BP",
+ allGenes=geneList,
+ annot = annFUN.gene2GO,
+ gene2GO = geneID2GO)
Error in .local(.Object, ...) : allGenes must be a factor with 2 levels
What did I miss?
Thank you in advance.
> genesOfInterest <- rownames(tr@.Data[[1]])
> head(genesOfInterest)
[1] "sp0025247" "sp0025250" "sp0025268" "sp0025270" "sp0025282" "sp0056834"
> geneList <- factor(as.integer(grep(genesOfInterest, geneUniverse)))
Warning message:
In grep(genesOfInterest, geneUniverse) :
argument 'pattern' has length > 1 and only the first element will be used
> head(geneList)
factor(0)
Levels:
> names(geneList) <- geneUniverse
Error in names(geneList) <- geneUniverse :
'names' attribute [44620] must be the same length as the vector [0]
Your second option geneUniverse <- gsub("\\\"", "", geneUniverse)) will be used with grep together?
Indeed, apologies, grep doesn't work with multiple values.
The grep code should be:
indices <- c()
for (i in 1:length(genesOfInterest)) indices <- c(indices, grep(genesOfInterest[i], geneUniverse))
geneList <- factor(as.integer(indices))
The second option should work without grep.
I have got 'names' attribute [44620] must be the same length as the vector [33975]
> genesOfInterest <- rownames(tr@.Data[[1]])
> head(genesOfInterest)
[1] "sp0025247" "sp0025250" "sp0025268" "sp0025270" "sp0025282" "sp0056834"
> indices <- c()
> for (i in 1:length(genesOfInterest)) indices <- c(indices, grep(genesOfInterest[i], geneUniverse))
> geneList <- factor(as.integer(indices))
> head(geneList)
[1] 12064 12076 26933 18988 18992 18994
33975 Levels: 2 3 4 5 6 8 10 11 12 13 14 15 18 19 22 23 24 25 26 28 29 30 31 32 33 34 35 ... 44618
> names(geneList) <- geneUniverse
Error in names(geneList) <- geneUniverse :
'names' attribute [44620] must be the same length as the vector [33975]
I think that your command > names(geneList) <- geneUniverse should be
names(geneList) <- geneUniverse[indices]
Nevertheless, in your other thread, which appears to be a duplicate of this, didn't you say that the geneList vector needs to have P values and then names as the gene names?
I have followed this tutorial for topGO. Only thing which I changed was:
> genesOfInterest <- read.table("interestinggenes.txt",header=FALSE)
> genesOfInterest <- as.character(genesOfInterest$V1)
to
> genesOfInterest <- rownames(tr@.Data[[1]])
Do you have any idea how to fix the geneList?
Thank you in advance.
I see, so that function is indeed just looking for indices (and not values).
It just looks like your matching is not working.
For example,
"\"sp0000005\"" is not the same as "sp0000005"
a <- c("\"sp0000005\"")
b <- c("sp0000005")
a %in% b
[1] FALSE
b %in% a
[1] FALSE
...however, grep returns indices too and only looks for partial matches (which can be risky at times):
grep(b, a)
[1] 1
Dear Ric,
I think that you just need to change the following line
geneList <- factor(as.integer(geneUniverse %in% genesOfInterest))
to:
EDIT: see next answer thread below
The other option is to add a new line of code after geneUniverse <- names(geneID2GO) that ensures that your variables in geneUniverse and genesOfInterest have the same name:
geneUniverse <- gsub("\\\"", "", geneUniverse))
Log in to answer this question.
Are you sure that the line
geneList <- factor(as.integer(geneUniverse %in% genesOfInterest))is doing what you expect it to do? First converting to integers and then factors doesn't look right to me. I'm neither sure that the lookup with%in%is going to work as you'd expect. The%in%operator will just return indices, but not actual values.Your geneList variable appears to be just zeros, i.e., a single factor.
Happy to assist further