DeSeq2 error correction and experiment design

Question

[abseq]

Hi All,

I am using DeSeq2 for differential expression analysis. I have treated (3 replicates) and untreated (3 replicates). I have used htseq-count for read count. For DEG I am following below link.

But I am facing some error. Below is the error.

df_count_matrix <- data.frame(file_name = dd) %>% + mutate(file_cont = map(file_name,fread,data.table = F)) %>% + unnest() %>% + mutate(file_name = gsub(pattern=pattern,replacement="",file_name)) %>% + spread(key = file_name , value = V2) Error in .f(.x[[i]], ...) : input= must be a single character string containing a file name, a system command containing at least one space, a URL starting 'http[s]://', 'ftp[s]://' or 'file://', or, the input data itself containing at least one \n or \r

Kindly suggest me how to correct the error and how should I design treated vs untreated.

Many thanks in advance.

[abseq]

Thank you ATpoint for quick reply... :)

[abseq]

I have some doubt, as mentioned in the steps to use "airway" package in the steps for RangedSummarizedExperiment. When I am using that it is adding the example data from that package.

And another doubt is as I have 2 samples and 3 replicates each (Treated - sample1, sample4, sample5, Untreated - sample2, sample3, sample6), how i should collaps them.

Thank you.

[GenoMax]

Please use ADD REPLY/ADD COMMENT when responding to existing posts to keep threads logically organized. Submit Answer is only for new answers to the original question.

[abseq]

Thanks genomax for the suggestion and I will follow it.

[ATpoint]

If you copy/paste the code from the manual that shows the process with the example data, you'll get example data, that is kind of expected ;-) Modify the paths to your htseq files. Don't ever collapse replicates, they are necessary for variance estimation. Please spend quality time reading about the basics of differential analysis before continuing with your analysis e.g. https://peerj.com/preprints/27283/

[abseq]

Actually I tried to used the airway package with my data and I think it was not the correct way. I am doing in htseq path. Yes, you are correct I should spend some time to understand the analysis.

Thanks ATpoint.

Answer 1

[ATpoint]

There is a dedicated function for htseq input. See the manual. Towards the design, it is a standard group vs group, the manual covers that as well (1~group).