Question

results(dds) error: couldn't find results. you should first run DESeq()

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beslinail • 0

@d4a633b9

Last seen 9 months ago

Turkey

Enter the body of text here

Code should be placed in three backticks as shown below

untar("gdc_download_20220310_195149.593210.tar.gz", exdir = "gdc_glandF_cancer")
untar("clinical.cart.2022-03-10.tar.gz")
sample_sheet <- read_tsv("gdc_sample_sheet.2022-03-10.tsv")
clinical <- read_tsv('clinical.tsv')
sampleTablejoin <- inner_join(sample_sheet, clinical,
                              by = c("Case ID" = "case_submitter_id"))

sampleTable <- sampleTablejoin %>%
  dplyr::select(`Sample ID`, `File Name`, Sample_Type) %>%
  distinct(`Sample ID`, .keep_all = TRUE) %>%
  dplyr::rename(condition = Sample_Type)



sampleTable <- sampleTablejoin %>%
  mutate(`File Name` = paste0(`File ID`, "/", `File Name`)) %>%
  dplyr::select(`Sample ID`, `File Name`, Sample_Type) %>%
  distinct(`Sample ID`, .keep_all = TRUE) %>%
  dplyr::rename(condition = Sample_Type)


dds <- DESeqDataSetFromHTSeqCount(sampleTable = as.data.frame(sampleTable),
                                  directory = "gdc_glandF_cancer",
                                  design = ~condition)

vsd <- vst(dds)
plotPCA(vsd)


dds <- DESeq(dds)
resdf <- as.data.frame(results(dds, contrast = c("condition", "Metastatic", "Solid_Tissue_Normal")))

Error in h(simpleError(msg, call)) : 
  error in evaluating the argument 'x' in selecting a method for function 'as.data.frame': couldn't find results. you should first run DESeq()

# include your problematic code here with any corresponding output 
# please also include the results of running the following in an R session 

sessionInfo( )

RNASeqRData • 3.1k views

ADD COMMENT • link updated 2.1 years ago by Michael Love 41k • written 2.1 years ago by beslinail • 0

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Take it step by step :

resdf=results(dds, contrast = c("condition", "Metastatic", "Solid_Tissue_Normal"))
resdf=as.data.frame(resdf)

ADD REPLY • link 2.1 years ago Basti ▴ 760

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I am taking the same error. It took from me too much time. Any help? I am a new one on this issue.

Thanks

ADD REPLY • link 2.1 years ago beslinail • 0

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Can you check dds has been correctly processed ? Please give us the output of dds in your console

ADD REPLY • link 2.1 years ago Basti ▴ 760

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> dds <- DESeq(dds)
estimating size factors
  Note: levels of factors in the design contain characters other than
  letters, numbers, '_' and '.'. It is recommended (but not required) to use
  only letters, numbers, and delimiters '_' or '.', as these are safe characters
  for column names in R. [This is a message, not a warning or an error]
estimating dispersions
gene-wise dispersion estimates
mean-dispersion relationship
  Note: levels of factors in the design contain characters other than
  letters, numbers, '_' and '.'. It is recommended (but not required) to use
  only letters, numbers, and delimiters '_' or '.', as these are safe characters
  for column names in R. [This is a message, not a warning or an error]
final dispersion estimates
  Note: levels of factors in the design contain characters other than
  letters, numbers, '_' and '.'. It is recommended (but not required) to use
  only letters, numbers, and delimiters '_' or '.', as these are safe characters
  for column names in R. [This is a message, not a warning or an error]
fitting model and testing
Error in h(simpleError(msg, call)) : 
  error in evaluating the argument 'x' in selecting a method for function 'rowSums': cannot allocate vector of size 179.0 Mb
> resdf <- results(dds, contrast = c("condition", "Metastatic", "Solid_Tissue_Normal"))
Error in results(dds, contrast = c("condition", "Metastatic", "Solid_Tissue_Normal")) : 
  couldn't find results. you should first run DESeq()
> resdf <- as.data.frame(resdf)
Error in h(simpleError(msg, call)) : 
  error in evaluating the argument 'x' in selecting a method for function 'as.data.frame': object 'resdf' not found

ADD REPLY • link updated 2.1 years ago by Michael Love 41k • written 2.1 years ago by beslinail • 0

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The main error is here : "Error in h(simpleError(msg, call)) : error in evaluating the argument 'x' in selecting a method for function 'rowSums': cannot allocate vector of size 179.0 Mb " You ran out of memory so the dds object has not been created. You can try memory.limit(size=56000)

ADD REPLY • link 2.1 years ago Basti ▴ 760

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HOW can I try memory.limit(size=56000) ? Would you clarify it, please?

ADD REPLY • link 2.1 years ago beslinail • 0

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Copy-paste my code, run it in your console and rerun your script

ADD REPLY • link 2.1 years ago Basti ▴ 760

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It really worked. Thank you so much

ADD REPLY • link 2.1 years ago beslinail • 0

0

Entering edit mode

I am taking the same error. It took from me too much time. Any help? I am a new one on this issue.

Thanks

ADD REPLY • link 2.1 years ago beslinail • 0

0

Entering edit mode

I am taking the same error. It took from me too much time. Any help? I am a new one on this issue.

Thanks

ADD REPLY • link 2.1 years ago beslinail • 0

score 0 · Answer 1 · 2022-03-11

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Michael Love 41k

@mikelove

Last seen 9 minutes ago

United States

What is the dim(dds)? And how much memory do you have to work with this data?

ADD COMMENT • link 2.1 years ago Michael Love 41k

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total system memory is 4,061 MiB Windows system

ADD REPLY • link 2.1 years ago beslinail • 0

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4 Gb should be fine for most RNA-seq datasets, but what about my first question? Are you working with an especially large number of features or samples?

ADD REPLY • link 2.1 years ago Michael Love 41k

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MY raw data is nearly 100 MB and the data is HTSeq - Counts to calculate the DEG analysis. There are three groups Solid Tissue Normal, Metastatic, Primary Tumor. but only I compare two groups of samples.

ADD REPLY • link 2.1 years ago beslinail • 0

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Ok sorry, I don't know what else to say, I've asked twice now about the dimensions of the object, but you're not saying. I guess if the raw htseq files are 100 MB then it shouldn't be an issue either.

I recommend to work with a bioinformatics group to help solve the error then. Try to run the same analysis on a different machine, this helps to identify the problem. You can save the dds object with save(dds, file="dds.rda") and then load() on a different machine to run DESeq().

ADD REPLY • link 2.1 years ago Michael Love 41k

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dim(dds) [1] 60483 411

ADD REPLY • link 2.1 years ago beslinail • 0

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Recommend to start with:

tab <- table(dds$condition)
lower_n <- 0.25 * min(tab)
keep <- rowSums(counts(dds) >= 10) >= lower_n
table(keep)
dds <- dds[keep,]

This will remove the genes that have single digits counts for most samples. As you have 60,000 x 400 samples it's just using up extra space on your machine to keep those near 0 counts around in the dataset.

ADD REPLY • link 2.1 years ago Michael Love 41k