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  • #16
    If you have downloaded and unzipped the annotation from

    (example for hg19), you can just specify the local path to this file instead of 'hg19'.

    Comment


    • #17
      It would be nice to include example of report to see what types of metrics it can calculate. I am curious about this.

      Comment


      • #18
        Hi! I got it to work, the output looks correct! It's strange though I can see Step 3 and Step 5 as unsuccessful, but the output looks finished (includes list of poor and high covered genes).

        I really like the summary.



        java NGSrich -r /net/ngs/NGS.analysis/data/091204_ILLUMINA-075005_0000_42GHRAAXX/s_1.42GHRAAXX_LIB001_MIAPACA2_24877_FID000.J00001.alnRecal.sam -a /net/ngs/NGS.analysis/bin/annovar/humandb/hg18_refGene.txt -t /net/ngs/NGS.analysis/reference/500_Target_Regions.bed -T /dev/shm/
        test: /net/ngs/NGS.analysis/data/091204_ILLUMINA-075005_0000_42GHRAAXX/enrichment
        =======================1=======================
        >>> STEP 1: reducing files
        READS FILE:
        /net/ngs/NGS.analysis/data/110415_SN378_0070_B81MCAABXX/s_1.B81MCAABXX_LIB160_HT_26828_FID047_J00229.bfast.sam was reduced to /dev/shm/1305908202090/NGSrich_s_1.B81MCAABXX_LIB160_HT_26828_FID047_J00229.bfast_vieasncl6.20049.txt
        Reduced file /dev/shm/1305908202090/NGSrich_s_1.B81MCAABXX_LIB160_HT_26828_FID047_J00229.bfast_vieasncl6.20049.txt sorted

        GENOME ANNOTATION FILE:
        /dev/shm/1305908202090/hg18_refGene.genome reduced to /dev/shm/1305908202090/NGSrich_genome_vieasncl6.20049.txt

        TARGET REGIONS FILE:
        /dev/shm/1305908202090/1000plus_Target_Regions.target reduced to /dev/shm/1305908202090/NGSrich_target_vieasncl6.20049.txt
        Reduced file /dev/shm/1305908202090/NGSrich_target_vieasncl6.20049.txt sorted

        STEP 1 successfully completed

        =====================2=========================
        >>> STEP 2: computing target coverage data

        Starting computing target coverage files.
        Mean target coverage data computed (/net/ngs/NGS.analysis/data/110415_SN378_0070_B81MCAABXX/enrichment/s_1.B81MCAABXX_LIB160_HT_26828_FID047_J00229.bfast_enrichment.bed).
        Detailed base coverage computed (/dev/shm/1305908202090/coverage_vieasncl6.20049.txt).
        Coverage summary computed (/net/ngs/NGS.analysis/data/110415_SN378_0070_B81MCAABXX/enrichment/s_1.B81MCAABXX_LIB160_HT_26828_FID047_J00229.bfast_enrichment.xml).

        STEP 2 successfully completed

        =====================3=========================
        >>> STEP 3: evaluating enrichment files

        Detecting evaluation program
        "data" and "plots" subdirectories created and xml and bed file moved to the data subdirectory!
        Run evaluation program on the following arguments Writing HTML output ... ready.Finished.
        HTML FILE /net/ngs/NGS.analysis/data/110415_SN378_0070_B81MCAABXX/enrichment/s_1.B81MCAABXX_LIB160_HT_26828_FID047_J00229.bfast_enrichment.html not found378_0070_B81MCAABXX/enrichment/110415_enrichment.html
        Creating coverage barplot ... ready.Searching for poorly (<2x) and highly (>200x) covered genes ...AABXX_LIB160_HT_26828_FID047_J00229.bfast_enrichment.bed
        STEP 3 unsuccessfulbarplots ... ready.Creating coverage barplot ...CAABXX/enrichment
        Creating coverage pieplot ... ready.Preparing coverage barplots ...efGene.txt
        =====================4=========================eplot ...
        >>> STEP 4: computing targets wiggle datale ...
        R Ausgabe: Reading XML file ...
        Start computing target-based wiggle data
        Details File Name: /dev/shm/1305908202090/coverage_vieasncl6.20049.txt
        Output Dir: /net/ngs/NGS.analysis/data/110415_SN378_0070_B81MCAABXX/enrichment/data
        End of computing target-based wiggle data

        STEP 4 successfully completed.

        =====================5=========================
        >>> STEP 5: computing overall wiggle data

        Start computing overall wiggle data
        Align File Name: /dev/shm/1305908202090/NGSrich_s_1.B81MCAABXX_LIB160_HT_26828_FID047_J00229.bfast_vieasncl6.20049.txt
        Output Dir: /net/ngs/NGS.analysis/data/110415_SN378_0070_B81MCAABXX/enrichment/data
        /net/ngs/NGS.analysis/data/110415_SN378_0070_B81MCAABXX/enrichment/s_1.B81MCAABXX_LIB160_HT_26828_FID047_J00229.bfast_enrichment.wignot found

        STEP 5 unsuccessful

        ===============================================

        [1]+ Done java NGSrich -r /net/ngs/NGS.analysis/data/110415_SN378_0070_B81MCAABXX/s_1.B81MCAABXX_LIB160_HT_26828_FID047_J00229.bfast.sam -a /net/ngs/NGS.analysis/bin/annovar/humandb/hg18_refGene.txt -t /net/ngs/NGS.analysis/reference/1000plus_Target_Regions.bed -T /dev/shm/

        Comment


        • #19
          Hi,

          I am having a similar problem. It seems that the sam file and the database are fine but then I get this error

          java NGSrich -r /Data/Exp1/s_3_QC_remdup.sam -a hg19 -T /Data/Exp1/ -t /Data/Exp1/exomeplusv2.bed
          test: /Data/Exp1/enrichment
          =======================1=======================
          >>> STEP 1: reducing files

          READS FILE:
          /Data/Exp1/s_3_QC_remdup.sam was reduced to /Data/Exp1/1306423511007/NGSrich_s_3_QC_remdup_KDavid.26819.txt
          Reduced file /Data/Exp1/1306423511007/NGSrich_s_3_QC_remdup_KDavid.26819.txt sorted

          GENOME ANNOTATION FILE:
          /Data/Exp1/1306423511007/refGene.genome reduced to /Data/Exp1/1306423511007/NGSrich_genome_KDavid.26819.txt
          Exception in thread "main" java.lang.ArrayIndexOutOfBoundsException: Array index out of range: 1
          at java.util.Vector.get(Vector.java:721)
          at adapter.Adapter.field(Adapter.java:90)
          at adapter.TargetAdapter.start(TargetAdapter.java:168)
          at adapter.TargetAdapter.adapt(TargetAdapter.java:56)
          at Enrichment.reduceFiles(Enrichment.java:185)
          at NGSrich.main(NGSrich.java:91)

          This is a head result of my .bed file

          head /Data/Exp1/exomeplusv2.bed
          track:exome
          chr1 30275 30431
          chr1 69069 70029
          chr1 228233 228354
          chr1 228471 228711
          chr1 367647 368608
          chr1 470971 471330
          chr1 621084 622045
          chr1 741165 741285
          chr1 745438 745558

          Any ideas?

          Thanks,

          Dave

          Comment


          • #20
            i keep getting this error after ~20mins. Any idea?

            Thanks,
            Jason


            java NGSrich -r 32815T_GATKrealigned_duplicates_marked.sam -g hg19 -t 0247401_D_BED_20090724_hg19.bed -T ngsrich_tmp -o ngsrich_out


            =======================1=======================
            >>> STEP 1: reducing files

            READS FILE:
            /mnt/Storage/All_Users/lij/testNGSrich/32815T_GATKrealigned_duplicates_marked.sam was reduced to /mnt/Storage/All_Users/lij/testNGSrich/ngsrich_tmp/1306393150286/NGSrich_32815T_GATKrealigned_duplicates_marked_pmc-bioinf02.14020.txt
            Reduced file /mnt/Storage/All_Users/lij/testNGSrich/ngsrich_tmp/1306393150286/NGSrich_32815T_GATKrealigned_duplicates_marked_pmc-bioinf02.14020.txt sorted


            java.io.FileNotFoundException: /mnt/Storage/All_Users/lij/testNGSrich/ngsrich_tmp/1306393150286/refGene.genome (No such file or directory)
            at java.io.FileInputStream.open(Native Method)
            at java.io.FileInputStream.<init>(FileInputStream.java:106)
            at java.util.Scanner.<init>(Scanner.java:636)
            at adapter.GenomeAdapter.adapt(GenomeAdapter.java:22)
            at Enrichment.reduceFiles(Enrichment.java:181)
            at NGSrich.main(NGSrich.java:91)
            Exception in thread "main" java.lang.NullPointerException
            at adapter.GenomeAdapter.adapt(GenomeAdapter.java:33)
            at Enrichment.reduceFiles(Enrichment.java:181)
            at NGSrich.main(NGSrich.java:91)

            Comment


            • #21
              Hi Peter,

              This looks like a very useful tool! Do you have plans to allow the input file to be in BAM rather than SAM format? Also, is it possible to include the median coverage along with the mean coverage?

              Thanks,
              Pete

              Comment


              • #22
                Hi,

                Originally posted by dnusol View Post
                Hi,

                I am having a similar problem. It seems that the sam file and the database are fine but then I get this error

                java NGSrich -r /Data/Exp1/s_3_QC_remdup.sam -a hg19 -T /Data/Exp1/ -t /Data/Exp1/exomeplusv2.bed
                test: /Data/Exp1/enrichment
                =======================1=======================
                >>> STEP 1: reducing files

                READS FILE:
                /Data/Exp1/s_3_QC_remdup.sam was reduced to /Data/Exp1/1306423511007/NGSrich_s_3_QC_remdup_KDavid.26819.txt
                Reduced file /Data/Exp1/1306423511007/NGSrich_s_3_QC_remdup_KDavid.26819.txt sorted

                GENOME ANNOTATION FILE:
                /Data/Exp1/1306423511007/refGene.genome reduced to /Data/Exp1/1306423511007/NGSrich_genome_KDavid.26819.txt
                Exception in thread "main" java.lang.ArrayIndexOutOfBoundsException: Array index out of range: 1
                at java.util.Vector.get(Vector.java:721)
                at adapter.Adapter.field(Adapter.java:90)
                at adapter.TargetAdapter.start(TargetAdapter.java:168)
                at adapter.TargetAdapter.adapt(TargetAdapter.java:56)
                at Enrichment.reduceFiles(Enrichment.java:185)
                at NGSrich.main(NGSrich.java:91)

                This is a head result of my .bed file

                head /Data/Exp1/exomeplusv2.bed
                track:exome
                chr1 30275 30431
                chr1 69069 70029
                chr1 228233 228354
                chr1 228471 228711
                chr1 367647 368608
                chr1 470971 471330
                chr1 621084 622045
                chr1 741165 741285
                chr1 745438 745558

                Any ideas?

                Thanks,

                Dave
                You should just remove the first line of the target file because it doesn't have the expected form (ie "chrome start end"). In the next version we will probably allow comments within the files.

                Regards,
                @abdallah

                Comment


                • #23
                  Hi,

                  @dnusol
                  You should remove the header from your .bed-file because it causes the parse error, since the program expects only lines with the following format (chrom start end). We will maybe allow comments or headers in the next versions.

                  Regards,
                  @abdallah
                  Last edited by @abdallah; 05-31-2011, 11:17 AM.

                  Comment


                  • #24
                    Hey this looks like a great tool - can it only be used for samtools/bwa alignments? what about novoalign?

                    Comment


                    • #25
                      Hi,

                      NGSrich looks very useful, I've been searching for a tool like that for a long time now...

                      However, I've got some problems ( I use version 0.4.3). I did alignemnt using bwa and a reference sequence which has chromosomes ordered 1-22XYMT with no unplaced unocalized or alternative contigs. The graphs for the first chromosomes (chr1, chr10, chr11, chr12, chr13, chr14, chr16 and chr 17) but it seems to discard the rest of the data. I just get empty plots for the other chromosomes. I was thinking about using a reference file with the chromosomes in the order as stated in the Alignment.html file, but that doesn't work as we analyze our data using GATK (and GATK wants to have the reference sequence ordered...)

                      I started your program as: java NGSrich -r Alignment.sam -a hg19 -t refGene.bed -T /tmp

                      Is that a problem of the reference sequence or am I making something wrong.

                      Thank you for your help and the great tool.
                      P.S.: BAM Support would indeed facilitate use of your tool!

                      Comment


                      • #26
                        Hi
                        I tried to run NGSrich_0.4.3 on a SAM file generated using Novoalign using the command
                        java NGSrich -r ../../B6113_NMT5_Novoalign/novoalign_simple/sam/B6113_novoalign.sam -g mm9 -t ../../B6113_NMT5_Novoalign/capture_target.alpha.bed

                        I received the following error:

                        test: /usr/local/work/hickey/NGSrich_0.4.3/bin/../../B6113_NMT5_Novoalign/novoalign_simple/sam/enrichment
                        File /scratch/ccg-ngs/tmp/1308184671325/ not found
                        Program stopped.


                        Can you please help?

                        Comment


                        • #27
                          I got the same error. It worked when I specified another tmp folder using the -T command (in my case -T /tmp)

                          Comment


                          • #28
                            Originally posted by ulz_peter View Post
                            I got the same error. It worked when I specified another tmp folder using the -T command (in my case -T /tmp)
                            Thanks, got it running!

                            Comment


                            • #29
                              Does this error show up in the output BED file already? This is located in the 'data' subdirectory and probably has only '0.0' coverage values for the chromosomes you mention. Could you pls email this BED file to me?

                              Regards,
                              Peter


                              Originally posted by ulz_peter View Post
                              Hi,

                              NGSrich looks very useful, I've been searching for a tool like that for a long time now...

                              However, I've got some problems ( I use version 0.4.3). I did alignemnt using bwa and a reference sequence which has chromosomes ordered 1-22XYMT with no unplaced unocalized or alternative contigs. The graphs for the first chromosomes (chr1, chr10, chr11, chr12, chr13, chr14, chr16 and chr 17) but it seems to discard the rest of the data. I just get empty plots for the other chromosomes. I was thinking about using a reference file with the chromosomes in the order as stated in the Alignment.html file, but that doesn't work as we analyze our data using GATK (and GATK wants to have the reference sequence ordered...)

                              I started your program as: java NGSrich -r Alignment.sam -a hg19 -t refGene.bed -T /tmp

                              Is that a problem of the reference sequence or am I making something wrong.

                              Thank you for your help and the great tool.
                              P.S.: BAM Support would indeed facilitate use of your tool!

                              Comment


                              • #30
                                Yes, the error shows up in the bed file. All the chromosomes which do not show any bars in the histogram have 0.0 values. Could you send a PM with your E-mail adress?

                                Comment

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