cummeRbund error in loading cuffdiff gene file
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Entering edit mode
10.3 years ago
Adrian Pelin ★ 2.6k

Hello,

I am trying to load my cuffdiff genes tracking file into cummeRbund and I get an error:

> refCuffdiff="/shared/AdrianP/home112013/glomus/RNA-Meiosis/JGI_annot/diff-cross-solo"
> gtfFilePath="/shared/AdrianP/home112013/glomus/RNA-Meiosis/JGI_annot/merged/merged.gtf"

> genomePath="/shared/AdrianP/home112013/glomus/RNA-Meiosis/JGI_annot/Gloin1_AssemblyScaffolds_Repeatmasked.fa"
> cuff <- readCufflinks(dir=refCuffdiff,rebuild=T,gtfFile=gtfFilePath,genome=genomePath)
Creating database /shared/AdrianP/home112013/glomus/RNA-Meiosis/JGI_annot/diff-cross-solo/cuffData.db
Reading Run Info File /shared/AdrianP/home112013/glomus/RNA-Meiosis/JGI_annot/diff-cross-solo/run.info
Writing runInfo Table
Reading Read Group Info  /shared/AdrianP/home112013/glomus/RNA-Meiosis/JGI_annot/diff-cross-solo/read_groups.info
Writing replicates Table
Reading GTF file
Writing GTF features to 'features' table...
Reading /shared/AdrianP/home112013/glomus/RNA-Meiosis/JGI_annot/diff-cross-solo/genes.fpkm_tracking
Error in scan(file, what, nmax, sep, dec, quote, skip, nlines, na.strings, :
  line 1 did not have 17 elements
>

I did not edit the genes.fpkm_tracking, here is how it looks after head:

tracking_id    class_code    nearest_ref_id    gene_id    gene_short_name    tss_id    locus    length    coverage    Crossing_FPKM    Crossing_conf_lo    Crossing_conf_hi    Crossing_status    Solo_FPKM    Solo_conf_lo    Solo_conf_hi    Solo_status
XLOC_000001    -    -    XLOC_000001    fgenesh1_kg.1_#_2_#_remain_c6340,fgenesh1_kg.1_#_3_#_step3_c2667,fgenesh1_pg.1_#_1,gm1.1_g    TSS1,TSS2,TSS3    scaffold_1:622-2570    -    -    21.9001    0    51.4671    OK    19.3357    0    51.4078    OK
XLOC_000002    -    -    XLOC_000002    gm1.12_g    TSS4    scaffold_10:10-512    -    -    0    0    0    OK    0    0    0    OK
XLOC_000003    -    -    XLOC_000003    gm1.154_g    TSS5    scaffold_100:720-1182    -    -    0.0767179    0    0.39657    OK    0    0    0    OK
XLOC_000004    -    -    XLOC_000004    gm1.153_g    TSS6    scaffold_100:21-396    -    -    0    0    0    OK    0    0    0    OK
XLOC_000005    -    -    XLOC_000005    fgenesh1_pg.1000_#_1,gm1.1557_g    TSS7,TSS8    scaffold_1000:0-2489    -    -    2.43942    0    8.08898    OK    2.15353    0    6.67501    OK
XLOC_000006    -    -    XLOC_000006    gm1.1558_g    TSS9    scaffold_1000:3054-3550    -    -    2.65628    0    8.96984    OK    5.06294    0    16.4288    OK
XLOC_000007    -    -    XLOC_000007    MIX1_1_43,e_gw1.10000.1.1,gm1.14515_g    TSS10    scaffold_10000:64-1868    -    -    0    0    0    OK    0.0292999    0    0.34366    OK
XLOC_000008    -    -    XLOC_000008    gm1.14516_g    TSS11,TSS12    scaffold_10001:164-1237    -    -    0    0    0    OK    0    0    0    OK
XLOC_000009    -    -    XLOC_000009    CE19_2291,CE23_3268,CE26_2185,CE29_2185,CE31_910,CE32_2410,fgenesh1_kg.10002_#_1_#_remain_c4528,fgenesh1_kg.10002_#_2_#_ACTTGA_L001_R1_(paired)_contig_254,fgenesh1_kg.10002_#_3_#_step3_c2559,fgenesh1_pg.10002_#_1    TSS13,TSS14,TSS15,TSS16,TSS17    scaffold_10002:1191-5743    -    -    205.934    0    427.865    OK    137.679    0    294.519    OK
XLOC_000010    -    -    XLOC_000010    fgenesh1_kg.10002_#_4_#_ACTTGA_L001_R1_(paired)_contig_4434    TSS18,TSS19,TSS20    scaffold_10002:1191-5743    -    -    89.4953    0    389.998    OK    125.531    0    422.435    OK
XLOC_000011    -    -    XLOC_000011    CE37_98,gm1.14519_g    TSS21,TSS22,TSS23    scaffold_10003:56-3724    -    -    3.44577    0    9.32412    OK    4.41548    0    12.7158    OK
XLOC_000012    -    -    XLOC_000012    gm1.14520_g    TSS24    scaffold_10004:23-239    -    -    0.726845    0    1.9669    OK    1.33937    0    3.53072    OK
XLOC_000013    -    -    XLOC_000013    fgenesh1_pg.10005_#_1,gm1.14521_g    TSS25    scaffold_10005:192-1657    -    -    0    0    0    OK    0    0    0    OK
XLOC_000014    -    -    XLOC_000014    gm1.14522_g    TSS26    scaffold_10006:251-777    -    -    0    0    0    OK    0    0    0    OK
XLOC_000015    -    -    XLOC_000015    gm1.14523_g    TSS27    scaffold_10007:900-1003    -    -    0    0    0    OK    0    0    0    OK
XLOC_000016    -    -    XLOC_000016    gm1.14524_g    TSS28    scaffold_10008:25-990    -    -    0    0    0    OK    0    0    0    OK
XLOC_000017    -    -    XLOC_000017    fgenesh1_pg.10009_#_1,gm1.14525_g,gm1.14526_g    TSS29,TSS30,TSS31    scaffold_10009:1648-3030    -    -    0    0    0    OK    0    0    0    OK
XLOC_000018    -    -    XLOC_000018    CE52_327,fgenesh1_kg.1001_#_1_#_ACTTGA_L001_R1_(paired)_contig_2142,fgenesh1_kg.1001_#_2_#_remain_c11121    TSS32,TSS33,TSS34,TSS35    scaffold_1001:464-2744    -    -    50.3672    0    150.486    OK    23.201    0    63.7491    OK
XLOC_000019    -    -    XLOC_000019    CE56_391,fgenesh1_kg.10010_#_1_#_step3_c842,gm1.14527_g    TSS36,TSS37    scaffold_10010:1116-2040    -    -    752.921    0    2536.24    OK    1132.62    0    5135    OK

Does anyone have any idea what went wrong?

Adrian

cufflinks cummeRbund R • 3.5k views
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Entering edit mode

can you show your code that you used to read in the cuffdiff output?

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I have modified my post to show that.

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It does not seem that there's anything wrong with genes.fpkm_tracking... maybe the scan() function is not opening the right file?

You tried making sym-links to a directory (or putting all input files into 1 directory) and then running readCufflinks() in that directory?

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Entering edit mode

I tried the same commands on a different experiment and I was in the same directory when I invoked R and it worked loading the genes tracking file.

Might it be that scan is parsing the file in a wrong way?

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Are you sure your genes.fpkm_tracking is tab-delimited? I think it should be tab-delimited for the program to correctly read the file.

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The default output is tab delimited, and what I posted is also tab delimited. Maybe it parses some columns incorrectly? THe gene_short_name column has many commas.

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Entering edit mode
10.3 years ago
Adrian Pelin ★ 2.6k

Alright, this is resolved now!

A quick email from Loyal A. Goff, a cummeRbund dev. hinted at the root of the problem:

Hi Adrian,

Without being able to reproduce the error first hand, my best guess at this point is the '#' characters in your gtf file.  I think others have had issues in the past with '#' because R considers everything after this to be a comment.  Is there any way that these characters can be removed from the reference gtf and cuffdiff could be rerun?

LoyK

Column 5 has these issues, and so does the gtf file. A quick sed fixed the issue.

Adrian

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