' )
+ f = file(dataset.file_name,'w')
+ f.write("\n".join( rval ))
+ f.write('\n')
+ f.close()
+
+ def set_peek( self, dataset, is_multi_byte=False ):
+ if not dataset.dataset.purged:
+ dataset.peek = "RSEM Reference (%s)" % ( dataset.metadata.reference_name )
+ dataset.blurb = "RSEM Reference (%s)" % ( dataset.metadata.reference_name )
+ else:
+ dataset.peek = 'RSEM Reference (%s) does not exist' % ( dataset.metadata.reference_name )
+ dataset.blurb = 'RSEM Reference (%s) purged from disk' % ( dataset.metadata.reference_name )
+
+ def display_peek( self, dataset ):
+ try:
+ return dataset.peek
+ except:
+ return "RSEM Reference"
+
+ def set_meta( self, dataset, overwrite = True, **kwd ):
+ """
+ Expecting files:
+ extra_files_path/.grp
+ extra_files_path/.ti
+ extra_files_path/.seq
+ extra_files_path/.transcripts.fa
+ Optionally includes files:
+ extra_files_path/.chrlist
+ extra_files_path/.idx.fa
+ extra_files_path/.4.ebwt
+ extra_files_path/.3.ebwt
+ extra_files_path/.2.ebwt
+ extra_files_path/.1.ebwt
+ extra_files_path/.rev.2.ebwt
+ extra_files_path/.rev.1.ebwt
+ """
+ log.info( "RSEM reference set_meta %s %s" % (dataset,dataset.extra_files_path))
+ pat = '^(.*)\.grp$'
+ efp = dataset.extra_files_path
+ flist = os.listdir(efp)
+ for i,fname in enumerate(flist):
+ m = re.match(pat,fname)
+ if m:
+ dataset.metadata.reference_name = m.groups()[0]
+ break
+ self.regenerate_primary_file(dataset)
+
+
diff -r 000000000000 -r 64d45f959303 rsem_calculate_expression.xml
--- /dev/null Thu Jan 01 00:00:00 1970 +0000
+++ b/rsem_calculate_expression.xml Mon Nov 11 13:54:43 2013 -0500
@@ -0,0 +1,583 @@
+
+ RNA-Seq by Expectation-Maximization
+
+ rsem
+ samtools
+ bowtie
+
+
+ rsem-calculate-expression
+ --calc-ci $useci.ci
+ --fragment-length-mean $fraglenmean
+ --fragment-length-min $fraglenmin
+ --fragment-length-sd $fraglensd
+ --fragment-length-max $fraglenmax
+ --bowtie-e $bowtie_e
+ --bowtie-m $bowtie_m
+
+ #if $input.format=="fastq"
+ ## IF FASTQ AND SINGLE END READS (DEFAULTS)
+ #if $input.fastqmatepair.matepair=="single" #rsem-wrapper-1.1.17.pl --bam_genome $bam_genome --bamtype $bamtype
+ --seed-length $seedlength $input.fastq_select --estimate-rspd $rspd --forward-prob
+ $fprob -p $cpus --bowtie-n $bowtie_mis --output-genome-bam --single_fastq $singlefastq
+ --output $output --isoformfile $isoforms --bamfile $bam_res --log $log
+ --sampling-for-bam $sampling_for_bam --reference ${index.fields.path}
+ #end if
+ ## IF FASTQ AND PAIRED END READS (DEFAULTS)
+ #if $input.fastqmatepair.matepair=="paired" #rsem-wrapper-1.1.17.pl --bam_genome $bam_genome --bamtype $bamtype
+ --paired-end --seed-length $seedlength --estimate-rspd $rspd $input.fastq_select --forward-prob $fprob -p $cpus
+ --bowtie-n $bowtie_mis --output-genome-bam --fastq1 $fastq1 --fastq2 $fastq2 --output
+ $output --isoformfile $isoforms --bamfile $bam_res --log $log --sampling-for-bam
+ $sampling_for_bam --reference ${index.fields.path}
+ #end if
+ #end if
+ #if $input.format=="fasta"
+ ## IF FASTA AND SINGLE END READS (DEFAULTS)
+ #if $input.fastamatepair.matepair=="single" #rsem-wrapper-1.1.17.pl --bam_genome $bam_genome --bamtype $bamtype
+ --no-qualities --seed-length $seedlength --estimate-rspd $rspd --forward-prob $fprob -p $cpus --bowtie-n $bowtie_mis
+ --output-genome-bam --single_fasta $single_fasta --output $output --isoformfile
+ $isoforms --bamfile $bam_res --log $log --sampling-for-bam $sampling_for_bam --reference
+ ${index.fields.path}
+ #end if
+ ## IF FASTA AND PAIRED END READS (DEFAULTS)
+ #if $input.fastamatepair.matepair=="paired" #rsem-wrapper-1.1.17.pl --bam_genome $bam_genome --bamtype $bamtype
+ --no-qualities --paired-end --seed-length $seedlength --estimate-rspd $rspd --forward-prob $fprob -p $cpus
+ --bowtie-n $bowtie_mis --output-genome-bam --fasta1 $fasta1 --fasta2 $fasta2 --output
+ $output --isoformfile $isoforms --bamfile $bam_res --log $log --sampling-for-bam
+ $sampling_for_bam --reference ${index.fields.path}
+ #end if
+ #end if
+
+
+
+
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+ bamtype == "yes"
+
+
+
+
+
+
+
+RSEM HOME PAGE - http://deweylab.biostat.wisc.edu/rsem/
+
+NAME
+ rsem-calculate-expression
+
+SYNOPSIS
+ rsem-calculate-expression [options] upstream_read_file(s) reference_name sample_name
+ rsem-calculate-expression [options] --paired-end upstream_read_file/s downstream_read_file/s reference_name sample_name
+ rsem-calculate-expression [options] --sam/--bam [--paired-end] input reference_name sample_name
+
+ARGUMENTS
+ upstream_read_files/s
+ Comma-separated list of files containing single-end reads or
+ upstream reads for paired-end data. By default, these files are
+ assumed to be in FASTQ format. If the --no-qualities option is
+ specified, then FASTA format is expected.
+
+ downstream_read_file/s
+ Comma-separated list of files containing downstream reads which are
+ paired with the upstream reads. By default, these files are assumed
+ to be in FASTQ format. If the --no-qualities option is specified,
+ then FASTA format is expected.
+
+ input
+ SAM/BAM formatted input file. If "-" is specified for the filename,
+ SAM/BAM input is instead assumed to come from standard input. RSEM
+ requires all alignments of the same read group together. For
+ paired-end reads, RSEM also requires the two mates of any alignment
+ be adjacent. See Description section for how to make input file obey
+ RSEM's requirements.
+
+ reference_name
+ The name of the reference used. The user must have run
+ 'rsem-prepare-reference' with this reference_name before running
+ this program.
+
+ sample_name
+ The name of the sample analyzed. All output files are prefixed by
+ this name (e.g., sample_name.genes.results)
+
+OPTIONS
+
+ --paired-end
+ Input reads are paired-end reads. (Default: off)
+
+ --no-qualities
+ Input reads do not contain quality scores. (Default: off)
+
+ --strand-specific
+ The RNA-Seq protocol used to generate the reads is strand specific,
+ i.e., all (upstream) reads are derived from the forward strand. This
+ option is equivalent to --forward-prob=1.0. With this option set, if
+ RSEM runs the Bowtie aligner, the '--norc' Bowtie option will be
+ used, which disables alignment to the reverse strand of transcripts.
+ (Default: off)
+
+ --sam
+ Input file is in SAM format. (Default: off)
+
+ --bam
+ Input file is in BAM format. (Default: off)
+
+ --sam-header-info [file]
+ RSEM reads header information from input by default. If this option
+ is on, header information is read from the specified file. For the
+ format of the file, please see SAM official website. (Default: "")
+
+ -p/--num-threads [int]
+ Number of threads to use. Both Bowtie and expression estimation will
+ use this many threads. (Default: 1)
+
+ --no-bam-output
+ Do not output any BAM file. (Default: off)
+
+ --output-genome-bam
+ Generate a BAM file, 'sample_name.genome.bam', with alignments
+ mapped to genomic coordinates and annotated with their posterior
+ probabilities. In addition, RSEM will call samtools (included in
+ RSEM package) to sort and index the bam file.
+ 'sample_name.genome.sorted.bam' and
+ 'sample_name.genome.sorted.bam.bai' will be generated. (Default:
+ off)
+
+ --sampling-for-bam
+ When RSEM generates a BAM file, instead of outputing all alignments
+ a read has with their posterior probabilities, one alignment is
+ sampled and outputed according to the posterior probabilities. If
+ the sampling result is that the read comes from the "noise"
+ transcript, nothing is outputed. (Default: off)
+
+ --calc-ci
+ Calculate 95% credibility intervals and posterior mean estimates.
+ (Default: off)
+
+ --seed-length [int]
+ Seed length used by the read aligner. Providing the correct value is
+ important for RSEM. If RSEM runs Bowtie, it uses this value for
+ Bowtie's seed length parameter. Any read with its or at least one of
+ its mates' (for paired-end reads) length less than this value will
+ be ignored. If the references are not added poly(A) tails, the
+ minimum allowed value is 5, otherwise, the minimum allowed value is
+ 25. Note that this script will only check if the value less or equal than
+ 5 and give a warning message if the value less than 25 but greter or equal than
+ 5. (Default: 25)
+
+ --tag [string]
+ The name of the optional field used in the SAM input for identifying
+ a read with too many valid alignments. The field should have the
+ format [tagName]:i:[value], where a [value] bigger than 0 indicates
+ a read with too many alignments. (Default: "")
+
+ --bowtie-path [path]
+ The path to the bowtie executables. (Default: the path to the bowtie
+ executables is assumed to be in the user's PATH environment
+ variable)
+
+ --bowtie-n [int]
+ (Bowtie parameter) max # of mismatches in the seed. (Range: 0-3,
+ Default: 2)
+
+ --bowtie-e [int]
+ (Bowtie parameter) max sum of mismatch quality scores across the
+ alignment. (Default: 99999999)
+
+ --bowtie-m [int]
+ (Bowtie parameter) suppress all alignments for a read if greater then [int]
+ valid alignments exist. (Default: 200)
+
+ --bowtie-chunkmbs [int]
+ (Bowtie parameter) memory allocated for best first alignment
+ calculation (Default: 0 - use bowtie's default)
+
+ --phred33-quals
+ Input quality scores are encoded as Phred+33. (Default: on)
+
+ --phred64-quals
+ Input quality scores are encoded as Phred+64 (default for GA
+ Pipeline ver. less than 1.3). (Default: off)
+
+ --solexa-quals
+ Input quality scores are solexa encoded (from GA Pipeline ver. less
+ than 1.3). (Default: off)
+
+ --forward-prob [double]
+ Probability of generating a read from the forward strand of a
+ transcript. Set to 1 for a strand-specific protocol where all
+ (upstream) reads are derived from the forward strand, 0 for a
+ strand-specific protocol where all (upstream) read are derived from
+ the reverse strand, or 0.5 for a non-strand-specific protocol.
+ (Default: 0.5)
+
+ --fragment-length-min [int]
+ Minimum read/insert length allowed. This is also the value for the
+ bowtie -I option. (Default: 1)
+
+ --fragment-length-max [int]
+ Maximum read/insert length allowed. This is also the value for the
+ bowtie -X option. (Default: 1000)
+
+ --fragment-length-mean [double]
+ (single-end data only) The mean of the fragment length distribution,
+ which is assumed to be a Gaussian. (Default: -1, which disables use
+ of the fragment length distribution)
+
+ --fragment-length-sd [double]
+ (single-end data only) The standard deviation of the fragment length
+ distribution, which is assumed to be a Gaussian. (Default: 0, which
+ assumes that all fragments are of the same length, given by the
+ rounded value of --fragment-length-mean)
+
+ --estimate-rspd
+ Set this option if you want to estimate the read start position
+ distribution (RSPD) from data. Otherwise, RSEM will use a uniform
+ RSPD. (Default: off)
+
+ --num-rspd-bins [int]
+ Number of bins in the RSPD. Only relevant when '--estimate-rspd' is
+ specified. Use of the default setting is recommended. (Default: 20)
+
+ --ci-memory [int]
+ Maximum size (in memory, MB) of the auxiliary buffer used for
+ computing credibility intervals (CI). Set it larger for a faster CI
+ calculation. However, leaving 2 GB memory free for other usage is
+ recommended. (Default: 1024)
+
+ --keep-intermediate-files
+ Keep temporary files generated by RSEM. RSEM creates a temporary
+ directory, 'sample_name.temp', into which it puts all intermediate
+ output files. If this directory already exists, RSEM overwrites all
+ files generated by previous RSEM runs inside of it. By default,
+ after RSEM finishes, the temporary directory is deleted. Set this
+ option to prevent the deletion of this directory and the
+ intermediate files inside of it. (Default: off)
+
+ --time
+ Output time consumed by each step of RSEM to 'sample_name.time'.
+ (Default: off)
+
+ -q/--quiet
+ Suppress the output of logging information. (Default: off)
+
+ -h/--help
+ Show help information.
+
+DESCRIPTION
+ In its default mode, this program aligns input reads against a reference
+ transcriptome with Bowtie and calculates expression values using the
+ alignments. RSEM assumes the data are single-end reads with quality
+ scores, unless the '--paired-end' or '--no-qualities' options are
+ specified. Users may use an alternative aligner by specifying one of the
+ --sam and --bam options, and providing an alignment file in the
+ specified format. However, users should make sure that they align
+ against the indices generated by 'rsem-prepare-reference' and the
+ alignment file satisfies the requirements mentioned in ARGUMENTS
+ section.
+
+ One simple way to make the alignment file satisfying RSEM's requirements
+ (assuming the aligner used put mates in a paired-end read adjacent) is
+ to use 'convert-sam-for-rsem' script. This script only accept SAM format
+ files as input. If a BAM format file is obtained, please use samtools to
+ convert it to a SAM file first. For example, if '/ref/mouse_125' is the
+ 'reference_name' and the SAM file is named 'input.sam', you can run the
+ following command:
+
+ convert-sam-for-rsem /ref/mouse_125 input.sam -o input_for_rsem.sam
+
+ For details, please refer to 'convert-sam-for-rsem's documentation page.
+
+ The SAM/BAM format RSEM uses is v1.4. However, it is compatible with old
+ SAM/BAM format. However, RSEM cannot recognize 0x100 in the FLAG field.
+ In addition, RSEM requires SEQ and QUAL are not '*'.
+
+ The user must run 'rsem-prepare-reference' with the appropriate
+ reference before using this program.
+
+ For single-end data, it is strongly recommended that the user provide
+ the fragment length distribution parameters (--fragment-length-mean and
+ --fragment-length-sd). For paired-end data, RSEM will automatically
+ learn a fragment length distribution from the data.
+
+ Please note that some of the default values for the Bowtie parameters
+ are not the same as those defined for Bowtie itself.
+
+ The temporary directory and all intermediate files will be removed when
+ RSEM finishes unless '--keep-intermediate-files' is specified.
+
+ With the '--calc-ci' option, 95% credibility intervals and posterior
+ mean estimates will be calculated in addition to maximum likelihood
+ estimates.
+
+OUTPUT
+ sample_name.genes.results
+ File containing gene level expression estimates. The format of each
+ line in this file is:
+
+ gene_id expected_counts tau_value [pmc_value tau_pme_value
+ tau_ci_lower_bound tau_ci_upper_bound] transcript_id_list
+
+ Fields are separated by the tab character. Fields within "[]" are
+ only presented if '--calc-ci' is set. pme stands for posterior mean
+ estimation. pmc stands for posterior mean counts. ci_lower_bound(l)
+ means the lower bound of the credibility intervals,
+ ci_upper_bound(u) means the upper bound of the credibility
+ intervals. So the credibility interval is [l, u].
+ 'transcript_id_list' is a space-separated list of transcript_ids
+ belonging to the gene. If no gene information is provided, this file
+ has the same content as 'sample_name.isoforms.results'.
+
+ sample_name.isoforms.results
+ File containing isoform level expression values. The format of each
+ line in this file is:
+
+ transcript_id expected_counts tau_value [pmc_value tau_pme_value
+ tau_ci_lower_bound tau_ci_upper_bound] gene_id
+
+ Fields are separated by the tab character. 'gene_id' is the gene_id
+ of the gene which this transcript belongs to. If no gene information
+ is provided, 'gene_id' and 'transcript_id' are the same.
+
+ sample_name.transcript.bam, sample_name.transcript.sorted.bam and
+ sample_name.transcript.sorted.bam.bai
+ Only generated when --no-bam-output is not specified.
+
+ 'sample_name.transcript.bam' is a BAM-formatted file of read
+ alignments in transcript coordinates. The MAPQ field of each
+ alignment is set to min(100, floor(-10 * log10(1.0 - w) + 0.5)),
+ where w is the posterior probability of that alignment being the
+ true mapping of a read. In addition, RSEM pads a new tag ZW:f:value,
+ where value is a single precision floating number representing the
+ posterior probability.
+
+ 'sample_name.transcript.sorted.bam' and
+ 'sample_name.transcript.sorted.bam.bai' are the sorted BAM file and
+ indices generated by samtools (included in RSEM package).
+
+ sample_name.genome.bam, sample_name.genome.sorted.bam and
+ sample_name.genome.sorted.bam.bai
+ Only generated when --no-bam-output is not specified and
+ --output-genome-bam is specified.
+
+ 'sample_name.genome.bam' is a BAM-formatted file of read alignments
+ in genomic coordinates. Alignments of reads that have identical
+ genomic coordinates (i.e., alignments to different isoforms that
+ share the same genomic region) are collapsed into one alignment. The
+ MAPQ field of each alignment is set to min(100, floor(-10 *
+ log10(1.0 - w) + 0.5)), where w is the posterior probability of that
+ alignment being the true mapping of a read. In addition, RSEM pads a
+ new tag ZW:f:value, where value is a single precision floating
+ number representing the posterior probability. If an alignment is
+ spliced, a XS:A:value tag is also added, where value is either '+'
+ or '-' indicating the strand of the transcript it aligns to.
+
+ 'sample_name.genome.sorted.bam' and
+ 'sample_name.genome.sorted.bam.bai' are the sorted BAM file and
+ indices generated by samtools (included in RSEM package).
+
+ sample_name.sam.gz
+ Only generated when the input files are raw reads instead of SAM/BAM
+ format files
+
+ It is the gzipped SAM output produced by bowtie aligner.
+
+ sample_name.time
+ Only generated when --time is specified.
+
+ It contains time (in seconds) consumed by aligning reads, estimating
+ expression levels and calculating credibility intervals.
+
+ sample_name.stat
+ This is a folder instead of a file. All model related statistics are
+ stored in this folder. Use 'rsem-plot-model' can generate plots
+ using this folder.
+
+EXAMPLES
+ Assume the path to the bowtie executables is in the user's PATH
+ environment variable. Reference files are under '/ref' with name
+ 'mouse_125'.
+
+ 1) '/data/mmliver.fq', single-end reads with quality scores. Quality
+ scores are encoded as for 'GA pipeline version >= 1.3'. We want to use 8
+ threads and generate a genome BAM file:
+
+ rsem-calculate-expression --phred64-quals \
+ -p 8 \
+ --output-genome-bam \
+ /data/mmliver.fq \
+ /ref/mouse_125 \
+ mmliver_single_quals
+
+ 2) '/data/mmliver_1.fq' and '/data/mmliver_2.fq', paired-end reads with
+ quality scores. Quality scores are in SANGER format. We want to use 8
+ threads and do not generate a genome BAM file:
+
+ rsem-calculate-expression -p 8 \
+ --paired-end \
+ /data/mmliver_1.fq \
+ /data/mmliver_2.fq \
+ /ref/mouse_125 \
+ mmliver_paired_end_quals
+
+ 3) '/data/mmliver.fa', single-end reads without quality scores. We want
+ to use 8 threads:
+
+ rsem-calculate-expression -p 8 \
+ --no-qualities \
+ /data/mmliver.fa \
+ /ref/mouse_125 \
+ mmliver_single_without_quals
+
+ 4) Data are the same as 1). We want to take a fragment length
+ distribution into consideration. We set the fragment length mean to 150
+ and the standard deviation to 35. In addition to a BAM file, we also
+ want to generate credibility intervals. We allow RSEM to use 1GB of
+ memory for CI calculation:
+
+ rsem-calculate-expression --bowtie-path /sw/bowtie \
+ --phred64-quals \
+ --fragment-length-mean 150.0 \
+ --fragment-length-sd 35.0 \
+ -p 8 \
+ --output-genome-bam \
+ --calc-ci \
+ --ci-memory 1024 \
+ /data/mmliver.fq \
+ /ref/mouse_125 \
+ mmliver_single_quals
+
+ 5) '/data/mmliver_paired_end_quals.bam', paired-end reads with quality
+ scores. We want to use 8 threads:
+
+ rsem-calculate-expression --paired-end \
+ --bam \
+ -p 8 \
+ /data/mmliver_paired_end_quals.bam \
+ /ref/mouse_125 \
+ mmliver_paired_end_quals
+
+
diff -r 000000000000 -r 64d45f959303 rsem_prepare_reference.xml
--- /dev/null Thu Jan 01 00:00:00 1970 +0000
+++ b/rsem_prepare_reference.xml Mon Nov 11 13:54:43 2013 -0500
@@ -0,0 +1,114 @@
+
+
+
+ rsem
+ bowtie
+
+
+ rsem-prepare-reference
+ #if $polya.polya_use == 'add':
+ #if $polya.polya_length:
+ --polyA-length $polya.polya_length
+ #end if
+ #elif $polya.polya_use == 'subset':
+ --no-polyA-subset $polya.no_polya_subset
+ #if $polya.polya_length:
+ --polyA-length $polya.polya_length
+ #end if
+ #elif $polya.polya_use == 'none':
+ --no-polyA
+ #end if
+ $ntog
+ #if $transcript_to_gene_map:
+ --transcript-to-gene-map $transcript_to_gene_map
+ #end if
+ #if $reference.ref_type == 'transcripts':
+ $reference.reference_fasta_file
+ #else:
+ --gtf $reference.gtf
+ $reference.reference_fasta_file
+ #end if
+ $reference_name
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+ Each line of should be of the form: gene_id transcript_id ( with the two fields separated by a tab character )
+ The map can be obtained from the UCSC table browser
+ group: Genes and Gene Prediction Tracks
+ table: knownIsoforms
+ Without a map:
+ If a reference genome and gtf is used, then RSEM uses the "gene_id" and "transcript_id" attributes in the GTF file.
+ Otherwise, RSEM assumes that each sequence in the reference sequence files is a separate gene.
+
+
+
+
+
+
+
+
+
+
+
+
+
+
+
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+
+
+
+
+
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+
+
+
+
+
+
+
+
+
+
+
+RSEM HOME PAGE - http://deweylab.biostat.wisc.edu/rsem/
+
+NAME
+ rsem-prepare-reference
+
+SYNOPSIS
+ rsem-prepare-reference [options] reference_fasta_file(s) reference_name
+
+DESCRIPTION
+ The rsem-prepare-reference program extracts/preprocesses the reference sequences and builds Bowtie indices using default parameters.
+ This program is used in conjunction with the 'rsem-calculate-expression' program.
+
+INPUTS
+
+
+
+
+
diff -r 000000000000 -r 64d45f959303 tool-data/rsem_indices.loc.sample
--- /dev/null Thu Jan 01 00:00:00 1970 +0000
+++ b/tool-data/rsem_indices.loc.sample Mon Nov 11 13:54:43 2013 -0500
@@ -0,0 +1,14 @@
+#This is a sample file distributed with Galaxy that enables tools
+#to use a directory of Bowtie indexed sequences data files. You will
+#need to create these data files and then create a bowtie_indices.loc
+#file similar to this one (store it in this directory) that points to
+#the directories in which those files are stored. The bowtie_indices.loc
+#file has this format (longer white space characters are TAB characters):
+#
+#
+#
+#So, for example, if you had hg18 indexed stored in
+#/depot/data2/galaxy/bowtie/hg18/,
+#then the bowtie_indices.loc entry would look like this:
+#
+#hg18 hg18 hg18 /depot/data2/galaxy/bowtie/hg18/hg18
diff -r 000000000000 -r 64d45f959303 tool_data_table_conf.xml.sample
--- /dev/null Thu Jan 01 00:00:00 1970 +0000
+++ b/tool_data_table_conf.xml.sample Mon Nov 11 13:54:43 2013 -0500
@@ -0,0 +1,8 @@
+
+
+