On May 1, 2013, at 1:43 PM, Iry Witham wrote:

I uploaded the bam file to galaxy main and get the same error. I also confirmed that the hg19.fa reference does exist in my cloudman instance. Any other ideas?

Thanks,

Iry

From: Dannon Baker <dannon.baker@gmail.com>
Date: Wednesday, May 1, 2013 12:38 PM
To: Iry Witham <iry.witham@jax.org>
Cc: "galaxy-dev@lists.bx.psu.edu" <galaxy-dev@lists.bx.psu.edu>
Subject: Re: [galaxy-dev] Installing FreeBayes via Toolshed: Tool dependency installation error

This one may be a little out of my wheelhouse, but did you verify that the reference it's trying to use is available? Does this same job work if you try it on galaxy main (if that's an option)?

reference=/mnt/galaxyIndices/genomes/Hsapiens/hg19/seq/hg19.fa

On Wed, May 1, 2013 at 8:41 AM, Iry Witham <Iry.Witham@jax.org> wrote:

Hi Dannon,

I was able to get cmake installed and then reinstalled Freebayes just fine. However, I now am getting the following error: unable to find FASTA index entry for '1'. The resulting files is:

Chrom Pos ID Ref Alt Qual Filter Info Format data

##fileformat=VCFv4.1

##fileDate=20130501

##source=freeBayes version 0.9.6

##reference=/mnt/galaxyIndices/genomes/Hsapiens/hg19/seq/hg19.fa

##phasing=none

##commandline="freebayes --bam localbam_0.bam --fasta-reference /mnt/galaxyIndices/genomes/Hsapiens/hg19/seq/hg19.fa --vcf /mnt/galaxyData/files/000/dataset_101.dat"

##INFO=<ID=NS,Number=1,Type=Integer,Description="Number of samples with data">

##INFO=<ID=DP,Number=1,Type=Integer,Description="Total read depth at the locus">

##INFO=<ID=AC,Number=A,Type=Integer,Description="Total number of alternate alleles in called genotypes">

##INFO=<ID=AN,Number=1,Type=Integer,Description="Total number of alleles in called genotypes">

##INFO=<ID=AF,Number=A,Type=Float,Description="Estimated allele frequency in the range (0,1]">

##INFO=<ID=RO,Number=1,Type=Integer,Description="Reference allele observations">

##INFO=<ID=AO,Number=A,Type=Integer,Description="Alternate allele observations">

##INFO=<ID=SRP,Number=1,Type=Float,Description="Strand balance probability for the reference allele: Phred-scaled upper-bounds estimate of the probability of observing the deviation between SRF and SRR given E(SRF/SRR) ~ 0.5, derived using Hoeffding's inequality">

##INFO=<ID=SAP,Number=A,Type=Float,Description="Strand balance probability for the alternate allele: Phred-scaled upper-bounds estimate of the probability of observing the deviation between SAF and SAR given E(SAF/SAR) ~ 0.5, derived using Hoeffding's inequality">

##INFO=<ID=AB,Number=A,Type=Float,Description="Allele balance at heterozygous sites: a number between 0 and 1 representing the ratio of reads showing the reference allele to all reads, considering only reads from individuals called as heterozygous">

##INFO=<ID=ABP,Number=A,Type=Float,Description="Allele balance probability at heterozygous sites: Phred-scaled upper-bounds estimate of the probability of observing the deviation between ABR and ABA given E(ABR/ABA) ~ 0.5, derived using Hoeffding's inequality">

##INFO=<ID=RUN,Number=A,Type=Integer,Description="Run length: the number of consecutive repeats of the alternate allele in the reference genome">

##INFO=<ID=RPP,Number=A,Type=Float,Description="Read Placement Probability: Phred-scaled upper-bounds estimate of the probability of observing the deviation between RPL and RPR given E(RPL/RPR) ~ 0.5, derived using Hoeffding's inequality">

##INFO=<ID=RPPR,Number=1,Type=Float,Description="Read Placement Probability for reference observations: Phred-scaled upper-bounds estimate of the probability of observing the deviation between RPL and RPR given E(RPL/RPR) ~ 0.5, derived using Hoeffding's inequality">

##INFO=<ID=EPP,Number=A,Type=Float,Description="End Placement Probability: Phred-scaled upper-bounds estimate of the probability of observing the deviation between EL and ER given E(EL/ER) ~ 0.5, derived using Hoeffding's inequality">

##INFO=<ID=EPPR,Number=1,Type=Float,Description="End Placement Probability for reference observations: Phred-scaled upper-bounds estimate of the probability of observing the deviation between EL and ER given E(EL/ER) ~ 0.5, derived using Hoeffding's inequality">

##INFO=<ID=DPRA,Number=A,Type=Float,Description="Alternate allele depth ratio. Ratio between depth in samples with each called alternate allele and those without.">

##INFO=<ID=XRM,Number=1,Type=Float,Description="Reference allele read mismatch rate: The rate of SNPs + MNPs + INDELs in reads supporting the reference allele.">

##INFO=<ID=XRS,Number=1,Type=Float,Description="Reference allele read SNP rate: The rate of per-base mismatches (SNPs + MNPs) in reads supporting the reference allele.">

##INFO=<ID=XRI,Number=1,Type=Float,Description="Reference allele read INDEL rate: The rate of INDELs (gaps) in reads supporting the reference allele.">

##INFO=<ID=XAM,Number=A,Type=Float,Description="Alternate allele read mismatch rate: The rate of SNPs + MNPs + INDELs in reads supporting the alternate allele, excluding the called variant.">

##INFO=<ID=XAS,Number=A,Type=Float,Description="Alternate allele read SNP rate: The rate of per-base mismatches (SNPs + MNPs) in reads supporting the alternate allele, excluding the called variant.">

##INFO=<ID=XAI,Number=A,Type=Float,Description="Alternate allele read INDEL rate: The rate of INDELs (gaps) in reads supporting the alternate allele, excluding the called variant.">

##INFO=<ID=ODDS,Number=1,Type=Float,Description="The log odds ratio of the best genotype combination to the second-best.">

##INFO=<ID=BVAR,Number=0,Type=Flag,Description="The best genotype combination in the posterior is variant (non homozygous).">

##INFO=<ID=CpG,Number=0,Type=Flag,Description="CpG site (either CpG, TpG or CpA)">

##INFO=<ID=TYPE,Number=A,Type=String,Description="The type of allele, either snp, mnp, ins, del, or complex.">

##INFO=<ID=CIGAR,Number=A,Type=String,Description="The extended CIGAR representation of each alternate allele, with the exception that '=' is replaced by 'M' to ease VCF parsing. Note that INDEL alleles do not have the first matched base (which is provided by default, per the spec) referred to by the CIGAR.">

##INFO=<ID=NUMALT,Number=1,Type=Integer,Description="Number of unique non-reference alleles in called genotypes at this position.">

##INFO=<ID=MEANALT,Number=A,Type=Float,Description="Mean number of unique non-reference allele observations per sample with the corresponding alternate alleles.">

##INFO=<ID=HWE,Number=1,Type=Float,Description="Phred-scaled discrete HWE prior probability of the genotyping across all samples.">

##INFO=<ID=LEN,Number=A,Type=Integer,Description="allele length">

##INFO=<ID=MQM,Number=A,Type=Float,Description="Mean mapping quality of observed alternate alleles">

##INFO=<ID=MQMR,Number=1,Type=Float,Description="Mean mapping quality of observed reference alleles">

##INFO=<ID=PAIRED,Number=A,Type=Float,Description="Proportion of observed alternate alleles which are supported by properly paired read fragments">

##INFO=<ID=PAIREDR,Number=1,Type=Float,Description="Proportion of observed reference alleles which are supported by properly paired read fragments">

##FORMAT=<ID=GT,Number=1,Type=String,Description="Genotype">

##FORMAT=<ID=GQ,Number=1,Type=Float,Description="Genotype Quality, the Phred-scaled marginal (or unconditional) probability of the called genotype">

##FORMAT=<ID=GL,Number=G,Type=Float,Description="Genotype Likelihood, log10-scaled likelihoods of the data given the called genotype for each possible genotype generated from the reference and alternate alleles given the sample ploidy">

##FORMAT=<ID=GLE,Number=1,Type=String,Description="Genotype Likelihood Explicit, same as GL, but with tags to indicate the specific genotype. For instance, 0^-75.22|1^-223.42|0/0^-323.03|1/0^-99.29|1/1^-802.53 represents both haploid and diploid genotype likilehoods in a biallelic context">

##FORMAT=<ID=DP,Number=1,Type=Integer,Description="Read Depth">

##FORMAT=<ID=RO,Number=1,Type=Integer,Description="Reference allele observation count">

##FORMAT=<ID=QR,Number=1,Type=Integer,Description="Sum of quality of the reference observations">

##FORMAT=<ID=AO,Number=A,Type=Integer,Description="Alternate allele observation count">

##FORMAT=<ID=QA,Number=A,Type=Integer,Description="Sum of quality of the alternate observations">

#CHROM POS ID REF ALT QUAL FILTER INFO FORMAT unknown

Iry

From: Dannon Baker <dannon.baker@gmail.com>
Date: Tuesday, April 30, 2013 1:06 PM
To: Iry Witham <iry.witham@jax.org>
Cc: "galaxy-dev@lists.bx.psu.edu" <galaxy-dev@lists.bx.psu.edu>
Subject: Re: [galaxy-dev] Installing FreeBayes via Toolshed: Tool dependency installation error

Looks like you're missing cmake, a dependency for building FreeBayes. If you're able to install that for your system (available in most package managers) you should be good to go.

-Dannon

On Tue, Apr 30, 2013 at 1:03 PM, Iry Witham <Iry.Witham@jax.org> wrote:

Hi All,

I am trying to get Freebayes installed on a cloud man instance from the galaxy toolshed. I have successfully installed the Samtools 0.1.18 package, but the freebayes package errors.

The Error is:

Tool shed repository: freebayes

Tool shed repository changeset revision: 28e422c589ac

Tool dependency status: Error

Tool dependency installation error: /bin/sh: cmake: not found

make[1]: *** [../bamtools/lib/libbamtools.a] Error 127

make: *** [all] Error 2

Tool dependency installation directory: /mnt/galaxyTools/tools/freebayes/0.9.6_9608597d12e127c847ae03aa03440ab63992fedf/devteam/freebayes/28e422c589ac

I get the following message after the error occurs:

This tool dependency is not installed correctly (see the Tool dependency installation error below). Choose Uninstall this tool dependency from the Repository Actions menu, correct problems if necessary, and try installing the dependency again.

I have followed this procedure and still have the same error. I have looked at the Installation.log file and this is what I found:

#############################################

git clone --recursive git://github.com/ekg/freebayes.git

STDOUT

Initialized empty Git repository in /mnt/galaxyData/tmp/tmpCUrxI7/freebayes/.git/

Submodule 'bamtools' (git://github.com/pezmaster31/bamtools.git) registered for path 'bamtools'

Submodule 'intervaltree' (git://github.com/ekg/intervaltree.git) registered for path 'intervaltree'

Submodule 'vcflib' (git://github.com/ekg/vcflib.git) registered for path 'vcflib'

Initialized empty Git repository in /mnt/galaxyData/tmp/tmpCUrxI7/freebayes/bamtools/.git/

Submodule path 'bamtools': checked out '3fe66b92028b60629826d827ee9058405fa7e4be'

Initialized empty Git repository in /mnt/galaxyData/tmp/tmpCUrxI7/freebayes/intervaltree/.git/

Submodule path 'intervaltree': checked out 'd151b487804861dc9f932e9f1fe4f8c499673cec'

Initialized empty Git repository in /mnt/galaxyData/tmp/tmpCUrxI7/freebayes/vcflib/.git/

Submodule path 'vcflib': checked out '480d96e16a552d36369dc9cb2b39ceedb260944a'

Submodule 'fastahack' (git://github.com/ekg/fastahack.git) registered for path 'fastahack'

Submodule 'fsom' (git://github.com/ekg/fsom.git) registered for path 'fsom'

Submodule 'intervaltree' (git://github.com/ekg/intervaltree.git) registered for path 'intervaltree'

Submodule 'multichoose' (git://github.com/ekg/multichoose.git) registered for path 'multichoose'

Submodule 'smithwaterman' (git://github.com/ekg/smithwaterman.git) registered for path 'smithwaterman'

Submodule 'tabixpp' (git://github.com/ekg/tabixpp.git) registered for path 'tabixpp'

Initialized empty Git repository in /mnt/galaxyData/tmp/tmpCUrxI7/freebayes/vcflib/fastahack/.git/

Submodule path 'fastahack': checked out '6acaf9875f2dbcdf86c9c690a8d188000e90d388'

Initialized empty Git repository in /mnt/galaxyData/tmp/tmpCUrxI7/freebayes/vcflib/fsom/.git/

Submodule path 'fsom': checked out '1b888f32c4bd571d1a0bea88d39cd7d7f23ea4b8'

Initialized empty Git repository in /mnt/galaxyData/tmp/tmpCUrxI7/freebayes/vcflib/intervaltree/.git/

Submodule path 'intervaltree': checked out '1290744283cef8076bb8a2968d4899b7228435f4'

Initialized empty Git repository in /mnt/galaxyData/tmp/tmpCUrxI7/freebayes/vcflib/multichoose/.git/

Submodule path 'multichoose': checked out '73d35daa18bf35729b9ba758041a9247a72484a5'

Initialized empty Git repository in /mnt/galaxyData/tmp/tmpCUrxI7/freebayes/vcflib/smithwaterman/.git/

Submodule path 'smithwaterman': checked out 'd9724f156c07cf16d00d251bebc20c6383eb6bde'

Initialized empty Git repository in /mnt/galaxyData/tmp/tmpCUrxI7/freebayes/vcflib/tabixpp/.git/

Submodule path 'tabixpp': checked out 'fbdde774adfc79ca75a65b4321b89b67cd81b921'

#############################################

#############################################

git clone --recursive git://github.com/ekg/freebayes.git

STDERR

#############################################

#############################################

git checkout 9608597d12e127c847ae03aa03440ab63992fedf

STDOUT

M bamtools

M vcflib

#############################################

#############################################

git checkout 9608597d12e127c847ae03aa03440ab63992fedf

STDERR

Note: checking out '9608597d12e127c847ae03aa03440ab63992fedf'.

You are in 'detached HEAD' state. You can look around, make experimental

changes and commit them, and you can discard any commits you make in this

state without impacting any branches by performing another checkout.

If you want to create a new branch to retain commits you create, you may

do so (now or later) by using -b with the checkout command again. Example:

git checkout -b new_branch_name

HEAD is now at 9608597... example pipeline script

#############################################

#############################################

git submodule update --recursive

STDOUT

Submodule path 'bamtools': checked out '8a5d6508d8fc52b25c9129037e44cc448bafebc7'

Submodule path 'vcflib': checked out '3f0d5e19596252b858a7f6eff1578e4295f549c8'

Submodule path 'fastahack': checked out 'd1bcd7a2d52ee0e2823884c8ede22ce228d3c060'

Submodule path 'smithwaterman': checked out '2882633a3b89f820d692a9b3316f4891dc3a1ff8'

#############################################

#############################################

git submodule update --recursive

STDERR

warning: unable to rmdir fsom: Directory not empty

#############################################

#############################################

make

STDOUT

cd src && make

make[1]: Entering directory `/mnt/galaxyData/tmp/tmpCUrxI7/freebayes/src'

cd ../bamtools && mkdir -p build && cd build && cmake .. && make

make[1]: Leaving directory `/mnt/galaxyData/tmp/tmpCUrxI7/freebayes/src'

#############################################

#############################################

make

STDERR

/bin/sh: cmake: not found

make[1]: *** [../bamtools/lib/libbamtools.a] Error 127

make: *** [all] Error 2

#############################################

Any ideas on how to resolve this?

Thanks,

Iry Witham

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Chrom	Pos	ID	Ref	Alt	Qual	Filter	Info	Format	data
##fileformat=VCFv4.1
##fileDate=20130501
##source=freeBayes version 0.9.6
##reference=/mnt/galaxyIndices/genomes/Hsapiens/hg19/seq/hg19.fa
##phasing=none
##commandline="freebayes --bam localbam_0.bam --fasta-reference /mnt/galaxyIndices/genomes/Hsapiens/hg19/seq/hg19.fa --vcf /mnt/galaxyData/files/000/dataset_101.dat"
##INFO=<ID=NS,Number=1,Type=Integer,Description="Number of samples with data">
##INFO=<ID=DP,Number=1,Type=Integer,Description="Total read depth at the locus">
##INFO=<ID=AC,Number=A,Type=Integer,Description="Total number of alternate alleles in called genotypes">
##INFO=<ID=AN,Number=1,Type=Integer,Description="Total number of alleles in called genotypes">
##INFO=<ID=AF,Number=A,Type=Float,Description="Estimated allele frequency in the range (0,1]">
##INFO=<ID=RO,Number=1,Type=Integer,Description="Reference allele observations">
##INFO=<ID=AO,Number=A,Type=Integer,Description="Alternate allele observations">
##INFO=<ID=SRP,Number=1,Type=Float,Description="Strand balance probability for the reference allele: Phred-scaled upper-bounds estimate of the probability of observing the deviation between SRF and SRR given E(SRF/SRR) ~ 0.5, derived using Hoeffding's inequality">
##INFO=<ID=SAP,Number=A,Type=Float,Description="Strand balance probability for the alternate allele: Phred-scaled upper-bounds estimate of the probability of observing the deviation between SAF and SAR given E(SAF/SAR) ~ 0.5, derived using Hoeffding's inequality">
##INFO=<ID=AB,Number=A,Type=Float,Description="Allele balance at heterozygous sites: a number between 0 and 1 representing the ratio of reads showing the reference allele to all reads, considering only reads from individuals called as heterozygous">
##INFO=<ID=ABP,Number=A,Type=Float,Description="Allele balance probability at heterozygous sites: Phred-scaled upper-bounds estimate of the probability of observing the deviation between ABR and ABA given E(ABR/ABA) ~ 0.5, derived using Hoeffding's inequality">
##INFO=<ID=RUN,Number=A,Type=Integer,Description="Run length: the number of consecutive repeats of the alternate allele in the reference genome">
##INFO=<ID=RPP,Number=A,Type=Float,Description="Read Placement Probability: Phred-scaled upper-bounds estimate of the probability of observing the deviation between RPL and RPR given E(RPL/RPR) ~ 0.5, derived using Hoeffding's inequality">
##INFO=<ID=RPPR,Number=1,Type=Float,Description="Read Placement Probability for reference observations: Phred-scaled upper-bounds estimate of the probability of observing the deviation between RPL and RPR given E(RPL/RPR) ~ 0.5, derived using Hoeffding's inequality">
##INFO=<ID=EPP,Number=A,Type=Float,Description="End Placement Probability: Phred-scaled upper-bounds estimate of the probability of observing the deviation between EL and ER given E(EL/ER) ~ 0.5, derived using Hoeffding's inequality">
##INFO=<ID=EPPR,Number=1,Type=Float,Description="End Placement Probability for reference observations: Phred-scaled upper-bounds estimate of the probability of observing the deviation between EL and ER given E(EL/ER) ~ 0.5, derived using Hoeffding's inequality">
##INFO=<ID=DPRA,Number=A,Type=Float,Description="Alternate allele depth ratio. Ratio between depth in samples with each called alternate allele and those without.">
##INFO=<ID=XRM,Number=1,Type=Float,Description="Reference allele read mismatch rate: The rate of SNPs + MNPs + INDELs in reads supporting the reference allele.">
##INFO=<ID=XRS,Number=1,Type=Float,Description="Reference allele read SNP rate: The rate of per-base mismatches (SNPs + MNPs) in reads supporting the reference allele.">
##INFO=<ID=XRI,Number=1,Type=Float,Description="Reference allele read INDEL rate: The rate of INDELs (gaps) in reads supporting the reference allele.">
##INFO=<ID=XAM,Number=A,Type=Float,Description="Alternate allele read mismatch rate: The rate of SNPs + MNPs + INDELs in reads supporting the alternate allele, excluding the called variant.">
##INFO=<ID=XAS,Number=A,Type=Float,Description="Alternate allele read SNP rate: The rate of per-base mismatches (SNPs + MNPs) in reads supporting the alternate allele, excluding the called variant.">
##INFO=<ID=XAI,Number=A,Type=Float,Description="Alternate allele read INDEL rate: The rate of INDELs (gaps) in reads supporting the alternate allele, excluding the called variant.">
##INFO=<ID=ODDS,Number=1,Type=Float,Description="The log odds ratio of the best genotype combination to the second-best.">
##INFO=<ID=BVAR,Number=0,Type=Flag,Description="The best genotype combination in the posterior is variant (non homozygous).">
##INFO=<ID=CpG,Number=0,Type=Flag,Description="CpG site (either CpG, TpG or CpA)">
##INFO=<ID=TYPE,Number=A,Type=String,Description="The type of allele, either snp, mnp, ins, del, or complex.">
##INFO=<ID=CIGAR,Number=A,Type=String,Description="The extended CIGAR representation of each alternate allele, with the exception that '=' is replaced by 'M' to ease VCF parsing. Note that INDEL alleles do not have the first matched base (which is provided by default, per the spec) referred to by the CIGAR.">
##INFO=<ID=NUMALT,Number=1,Type=Integer,Description="Number of unique non-reference alleles in called genotypes at this position.">
##INFO=<ID=MEANALT,Number=A,Type=Float,Description="Mean number of unique non-reference allele observations per sample with the corresponding alternate alleles.">
##INFO=<ID=HWE,Number=1,Type=Float,Description="Phred-scaled discrete HWE prior probability of the genotyping across all samples.">
##INFO=<ID=LEN,Number=A,Type=Integer,Description="allele length">
##INFO=<ID=MQM,Number=A,Type=Float,Description="Mean mapping quality of observed alternate alleles">
##INFO=<ID=MQMR,Number=1,Type=Float,Description="Mean mapping quality of observed reference alleles">
##INFO=<ID=PAIRED,Number=A,Type=Float,Description="Proportion of observed alternate alleles which are supported by properly paired read fragments">
##INFO=<ID=PAIREDR,Number=1,Type=Float,Description="Proportion of observed reference alleles which are supported by properly paired read fragments">
##FORMAT=<ID=GT,Number=1,Type=String,Description="Genotype">
##FORMAT=<ID=GQ,Number=1,Type=Float,Description="Genotype Quality, the Phred-scaled marginal (or unconditional) probability of the called genotype">
##FORMAT=<ID=GL,Number=G,Type=Float,Description="Genotype Likelihood, log10-scaled likelihoods of the data given the called genotype for each possible genotype generated from the reference and alternate alleles given the sample ploidy">
##FORMAT=<ID=GLE,Number=1,Type=String,Description="Genotype Likelihood Explicit, same as GL, but with tags to indicate the specific genotype. For instance, 0^-75.22\|1^-223.42\|0/0^-323.03\|1/0^-99.29\|1/1^-802.53 represents both haploid and diploid genotype likilehoods in a biallelic context">
##FORMAT=<ID=DP,Number=1,Type=Integer,Description="Read Depth">
##FORMAT=<ID=RO,Number=1,Type=Integer,Description="Reference allele observation count">
##FORMAT=<ID=QR,Number=1,Type=Integer,Description="Sum of quality of the reference observations">
##FORMAT=<ID=AO,Number=A,Type=Integer,Description="Alternate allele observation count">
##FORMAT=<ID=QA,Number=A,Type=Integer,Description="Sum of quality of the alternate observations">
#CHROM	POS	ID	REF	ALT	QUAL	FILTER	INFO	FORMAT	unknown