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Checking General-read_dir parameter ... error!

Issue #115 resolved
Rupinder Kaur created an issue

I'm working on Ubuntu18.04. I'm trying to run example. When i try to run example file. I got this error. I'm attaching the file.

Comments (27)

  2. Rupinder Kaur reporter

    No Sir, I haven't downloaded the files yet. But I will and try to run it again.

    Sir another problem I'm facing is in downloading the other data files like human_genom, bowtie index, mirDeep index etc.. The link that has provided in the documentation "http://miarmaseq.cbbio.es/download/..." is not working. Is there any other link where I can download these files.

  5. Rupinder Kaur reporter

    Sir, I didn't find url to download hg19_genome.tar.bz2. The link you have provided is only for index files and the link that is given in documentation is "http://miarmaseq.cbbio.es/download/Genome/hg19_genome.tar.bz2", that is expired.

  9. Rupinder Kaur reporter

    SIr, I'm trying to run miARma using this conf file.

    ;General parameters [General] ; type of analysis (miRNA, mRNA or circRNA) type=miRNA ; Folder for miRNA reads read_dir=/home/rupinder/ncbi/public/fastq ; label for the analsysis label=OSCC ; Folder where miARma has been instaled miARmaPath=/home/rupinder/NGS/cbbio-miarma-42e187a7f514 ; Folder to store results output_dir=/home/rupinder/NGS/results ; organism used organism=human ; Number of process to run at the same time threads=4 ; Whether the data is from a strand-specific assay (yes, no or reverse, yes by default) for featureCounts analysis strand=yes stats_file=/home/rupinder/NGS/results/mirna_stat.log logfile=/home/rupinder/NGS/results/miARma_logfile.log

    [Quality] prefix=Both

    [Adapter] ; Specific software to remove the adapter from the sequences adaptersoft=CutAdapt ; ; #Minimun length of the sequence read to keep with Cutadapt and Reaper Software min=18 ; ; #Maximun length of the sequence read to use with Cutadapt Software max=26 ; ; #Minimun quality of the sequence read to use with Cutadapt Software min_quality=25 ; #Number of adapter predictions to show adaptpredictionnumber=12

    [Aligner] ; Aligner (Bowtie1, Bowtie2, BWA or miRDeep) aligner=Bowtie1 ; Bowtie 2 index bowtie1index=/home/rupinder/NGS/cbbio-miarma-42e187a7f514/Index_bowtie1_hg19/Genomes/Indexes/bowtie1/human/bw1_homo_sapiens19

    [ReadCount] #GFF file used to calculate the number of reads in featureCounts analysis database=/home/rupinder/NGS/cbbio-miarma-42e187a7f514/Examples_miARma_miRNAs/Examples/basic_examples/miRNAs/data/miRBase_Annotation_20_for_hsa_mature_miRNA.gtf ;GFF attribute to be used as feature ID (default: gene_id) for featureCounts analysis seqid=transcript_id ;Feature type (3rd column in GFF file) to be used, all features of other type are ignored (default:exon) for featureCounts analysis featuretype=miRNA

    [DEAnalysis] ; Complete path of the target file. targetfile=/home/rupinder/NGS/cbbio-miarma-42e187a7f514/target.txt ; Path of the contrast file. contrastfile=/home/rupinder/NGS/cbbio-miarma-42e187a7f514/contrast.txt #This value refers to filter processing in the reads (Should be "yes" or "no"). filter=yes ;Specific software to perform the Differential Expression Analysis (Allowed values: edger, noiseq or edger-noiseq) desoft=EdgeR ;Provide a file with normalized reads cpm=yes ;Provide a file with RPKM values rpkm=yes

    [TargetPrediction]

    It works normally till quality check but at adapter removal step it shows some error. error.png.

  10. Eduardo Andres Leon

    It seems that you are using Minion to predict an adapter sequences. In that case, we check that the predicted sequence is not a sequences contained in the organism, to do that we use the Blat utility by the UCSC. As I see (other people report that), your network does not allow a connection to the UCSC. I recommend you to ask your admin to open connection to UCSC

  11. Rupinder Kaur reporter

    Hello SIr again, I'm getting an error at differential analysis step. I already have followed the #Issue63 and #Issue55 to resolve this error but still i'm getting this error again. error_edge.png Thanks in advance! :-)

  12. Eduardo Andres Leon

    Fine! you are almost done.

    Could you send me both log files ? (miarma_log and miARma_stats ???) E

  15. Eduardo Andres Leon

    Please do the following:

    bash$>cd /home/rupinder/NGS/cbbio-miarma-42e187a7f514
bash$>R
R>source("/home/rupinder/NGS/cbbio-miarma-42e187a7f514/lib/CbBio/RNASeq/R-Scripts/DE_EdgeR.R")
R>setwd("/home/rupinder/NGS/results/Readcount_results")
R> DE_EdgeR(projectdir="/home/rupinder/NGS/results/EdgeR_results",dir="/home/rupinder/NGS/results/Readcount_results", file="/home/rupinder/NGS/results/Readcount_results/cut_bw1-ReadCount.tab", targetfile="/home/rupinder/NGS/cbbio-miarma-42e187a7f514/target.txt", label="OSCC_cut_bw1", contrastfile="/home/rupinder/NGS/cbbio-miarma-42e187a7f514/contrast.txt", filter="yes", rpkm="yes", file_size="/home/rupinder/NGS/results/Readcount_results/cut_bw1-Size.tab", cpm="yes")

    You will see an error coming from the last command

  18. Rupinder Kaur reporter

    DE_EdgeR(projectdir="/home/rupinder/NGS/results/EdgeR_results",dir="/home/rupinder/NGS/results/Readcount_results", file="/home/rupinder/NGS/results/Readcount_results/cut_bw1-ReadCount.tab", targetfile="/home/rupinder/NGS/cbbio-miarma-42e187a7f514/target.txt", label="OSCC_cut_bw1", contrastfile="/home/rupinder/NGS/cbbio-miarma-42e187a7f514/contrast.txt", filter="yes", rpkm="yes", file_size="/home/rupinder/NGS/results/Readcount_results/cut_bw1-Size.tab", cpm="yes") Loading required package: edgeR Loading required package: limma 2018-06-26 15:17:16 Starting Differential expression analysis with EdgeR of OSCC_cut_bw1 [2018-06-26 15:17:16] Calculating normalized CPMs [2018-06-26 15:17:16] Calculating RPKMs [2018-06-26 15:17:16] RPKMs done! Error in .compressDispersions(y, dispersion) : dispersions must be finite non-negative values In addition: Warning message: In estimateCommonDisp.DGEList(dgenorm) : There is no replication, setting dispersion to NA.

  23. Eduardo Andres Leon

    As I see you have 3 samples and 0 replicates from each sample.

    To do a differential expression analysis you need 3 Norm, 3 OL and 3 OSCC samples (with 2 biological replicates could work).

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