Question: biomaRt correct annotation but...
0
gravatar for Mozart
2.4 years ago by
Mozart140
Mozart140 wrote:

Hi everyone, I am a bit confused about a painful step of my pipeline using Sleuth. I am trying to move from the transcript level to the gene level and that's why I am using biomaRt (?).

devtools::install_github('pachterlab/sleuth', ref = 'devel', force=TRUE)
mart <- biomaRt::useMart(biomart = "ensembl", dataset = "mmusculus_gene_ensembl", host = 'ensembl.org')
mart
t2g <- biomaRt::getBM(attributes = c("ensembl_transcript_id", "ensembl_gene_id",
                                 "external_gene_name"), mart = mart)
t2g <- dplyr::rename(t2g, target_id = ensembl_transcript_id,
                 ens_gene = ensembl_gene_id, ext_gene = external_gene_name)
so <- sleuth_prep(s2c, target_mapping = t2g)

at this point the output is the following one:

so <- sleuth_prep(s2c, target_mapping = t2g)
reading in kallisto results
dropping unused factor levels
......
normalizing est_counts
47250 targets passed the filter
normalizing tpm
merging in metadata
summarizing bootstraps
......
Warning messages:
1: In check_num_cores(num_cores) :
  It appears that you are running Sleuth from within Rstudio.
Because of concerns with forking processes from a GUI, 'num_cores' is being set to 1.
If you wish to take advantage of multiple cores, please consider running sleuth from the command line.
2: In check_target_mapping(tmp_names, target_mapping) :
  intersection between target_id from kallisto runs and the target_mapping is empty. attempted to fix problem by removing .N from target_id, then merging back into target_mapping. please check obj$target_mapping to ensure this new mapping is correct.

I can't understand the problem because I am pretty sure I have already created a table with the specific gene names and they match!

                target_id           ens_gene         ext_gene
333    ENSMUST00000148678 ENSMUSG00000020225           Tmbim4

Help?(!)

PS: please pardon my ignorance in term of bioinformatic jargon

rna-seq • 999 views
ADD COMMENTlink modified 2.4 years ago by haroldpimentel90 • written 2.4 years ago by Mozart140
3
gravatar for haroldpimentel
2.4 years ago by
haroldpimentel90 wrote:

Hey there,

Do you get results after running a test?

Basically, there is some weirdness between some of the ensembl annotations. Your transcriptome probably says something like 'ENSMUST00000148678.2' for that specific transcript. When we try to merge that with the target_mapping, it ends up being empty because the target_id is 'ENSMUST00000148678' (note the missing .2). It's basically just telling you that we removed .N and it seems to have fixed it. When we do that and there is no intersection mapping still, then the program will completely error.

If you still get results after running a test and calling sleuth_results(), then everything should be fine.

Best,

Harold

ADD COMMENTlink written 2.4 years ago by haroldpimentel90

Thanks a lot Harold, actually everything should be fine even if I am getting the error.

ADD REPLYlink written 2.4 years ago by Mozart140
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