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1 <tool id="rsem_calculate_expression" name="RSEM calculate expression" version="1.1.17">
2 <description>RNA-Seq by Expectation-Maximization</description>
3 <requirements>
4 <requirement type="package" version="1.1.17">rsem</requirement>
5 <requirement type="package" version="0.1.19">samtools</requirement>
6 <requirement type="package" version="1.0.0">bowtie</requirement>
7 </requirements>
8 <command>
9 rsem-calculate-expression
10 ## --tag string
11 #if $seedlength:
12 --seed-length $seedlength
13 #end if
14 --forward-prob $forward_prob
15 #if $rsem_options.fullparams == 'fullset':
16 ## Fragment info
17 #if $rsem_options.fragment_length_mean:
18 --fragment-length-mean $rsem_options.fragment_length_mean
19 #end if
20 #if $rsem_options.fragment_length_min:
21 --fragment-length-min $rsem_options.fragment_length_min
22 #end if
23 #if $rsem_options.fragment_length_sd:
24 --fragment-length-sd $rsem_options.fragment_length_sd
25 #end if
26 #if $rsem_options.fragment_length_max:
27 --fragment-length-max $rsem_options.fragment_length_max
28 #end if
29 ## RSPD
30 #if $rsem_options.rspd.estimate == 'yes':
31 --estimate-rspd
32 #if $rsem_options.rspd.num_rspd_bins:
33 --num-rspd-bins $rsem_options.rspd.num_rspd_bins
34 #end if
35 #end if
36 ## Calculate 95% credibility intervals and posterior mean estimates.
37 #if $rsem_options.useci.ci == 'yes':
38 --calc-ci
39 #if $rsem_options.useci.cimem:
40 --ci-memory $rsem_options.useci.cimem
41 #end if
42 #end if
43 #end if
44 ## --num-threads $GALAXY_SLOTS
45 #if $input.format != 'bam' and $input.bowtie_options.fullparams == 'fullset':
46 ## Bowtie params
47 #if $bowtie_options.bowtie_e:
48 --bowtie-e $bowtie_options.bowtie_e
49 #end if
50 #if $bowtie_options.bowtie_m:
51 --bowtie-m $bowtie_options.bowtie_m
52 #end if
53 #if $bowtie_options.bowtie_n:
54 --bowtie-n $bowtie_options.bowtie_n
55 #end if
56 #end if
57 ## Outputs
58 #if $rsem_outputs.result_bams == 'none':
59 --no-bam-output
60 #else
61 #if $rsem_outputs.result_bams == 'both':
62 --output-genome-bam
63 #end if
64 $rsem_outputs.sampling_for_bam
65 #end if
66 ## Input data
67 #if $input.format=="fastq"
68 $input.fastq_select
69 #if $input.fastq.matepair=="single":
70 $input.fastq.singlefastq
71 #elif $input.fastq.matepair=="paired":
72 --paired-end
73 $input.fastq.fastq1
74 $input.fastq.fastq2
75 #end if
76 #elif $input.format=="fasta"
77 --no-qualities
78 #if $input.fasta.matepair=="single":
79 $input.fasta.singlefasta
80 #elif $input.fasta.matepair=="paired":
81 --paired-end
82 $input.fasta.fasta1
83 $input.fasta.fasta2
84 #end if
85 #elif $input.format=="sam"
86 #if $input.matepair=="paired":
87 --paired-end
88 #end if
89 #if $input.rsem_sam._extension == 'sam':
90 --sam
91 #elif $input.rsem_sam._extension == 'bam':
92 --bam
93 #end if
94 $input.rsem_sam
95 #end if
96 ## RSEM reference
97 #if $reference.refSrc == 'history':
98 ${reference.rsem_ref.extra_files_path}/${reference.rsem_ref.metadata.reference_name}
99 #elif $reference.refSrc == 'cached':
100 ${reference.index.fields.path}
101 #end if
102 ## sample_name: use a hard coded name so we can pull out galaxy outputs
103 rsem_output
104 ## direct output into logfile
105 > $log
106 </command>
107 <macros>
108 <macro name="rsem_options">
109 <param name="seedlength" type="integer" value="25" optional="true" label="Seed length used by the read aligner" help="Providing the correct value for this parameter is important for RSEM's accuracy if the data are single-end reads. RSEM uses this value for Bowtie's seed length parameter. The minimum value is 25. (Default:25)">
110 </param>
111 <param name="forward_prob" type="select" label="Is the library strand specific?">
112 <option value="0.5" selected="true">No</option>
113 <option value="1">Yes, the reads (or first reads from paired-end libraries) are only in the forward orientation</option>
114 <option value="0">Yes, the reads (or first reads from paired-end libraries) are only in the reverse orientation</option>
115 </param>
116 <conditional name="rsem_options">
117 <param name="fullparams" type="select" label="Additional RSEM options">
118 <option value="default">Use RSEM Defaults</option>
119 <option value="fullset">Set Additional RSEM Options</option>
120 </param>
121 <when value="default"/>
122 <when value="fullset">
123 <param name="fragment_length_min" type="integer" value="1" optional="true" label="Minimum read/insert length." help=" This is also the value for the bowtie -I option">
124 <validator type="in_range" message="0 or greater" min="0" />
125 </param>
126 <param name="fragment_length_max" type="integer" value="1000" optional="true" label="Maximum read/insert length." help=" This is also the value for the bowtie -X option">
127 <validator type="in_range" message="0 or greater" min="0" max="1000000"/>
128 </param>
129 <param name="fragment_length_mean" type="float" value="" optional="true" label="Fragment length mean (single-end data only)" help="The mean of the fragment length distribution, which is assumed to be a Gaussian. (Default: -1, which disables use of the fragment length distribution)">
130 </param>
131 <param name="fragment_length_sd" type="float" value="" optional="true" label="The standard deviation of the fragment length distribution (single-end data only)" help="Default 0, which assumes that all fragments are of the same length, given by the rounded value of fragment length mean. ">
132 </param>
133 <conditional name="rspd">
134 <param name="estimate" type="select" lanel="Read Start Position Distribution (RSPD)"
135 help="Set this option if you want to estimate the read start position distribution (RSPD) from data. Otherwise, RSEM will use a uniform RSPD.">
136 <option value="no" selected="true">Use a uniform RSPD</option>
137 <option value="yes">Estimate and correct for a non-uniform RSPD</option>
138 </param>
139 <when value="no"/>
140 <when value="yes">
141 <param name="num_rspd_bins" type="integer" value="20" optional="true" label="Number of bins in the RSPD." help="Use of the default setting of 20 is recommended.">
142 <validator type="in_range" message="" min="0" max="100"/>
143 </param>
144 </when>
145 </conditional>
146 <conditional name="useci">
147 <param name="ci" type="select" label="Calculate 95% Credibility Intervals">
148 <option value="no" selected="true">no</option>
149 <option value="yes">yes</option>
150 </param>
151 <when value="no"/>
152 <when value="yes">
153 <param name="cimem" size="4" type="text" value="1024" label="Amount of memory in (MB) for computing CI" />
154 </when>
155 </conditional>
156 </when>
157 </conditional>
158 </macro>
159 <macro name="bowtie_options">
160 <conditional name="bowtie_options">
161 <param name="fullparams" type="select" label="bowtie settings">
162 <option value="default">use bowtie defaults</option>
163 <option value="fullset">set bowtie options</option>
164 </param>
165 <when value="default"/>
166 <when value="fullset">
167 <param name="bowtie_n" type="integer" value="2" optional="true" label="Bowtie mismatches" help="Bowtie parameter max # of mismatches in the seed. (Range: 0-3, Default: 2) ">
168 <validator type="in_range" message="max # of mismatches in the seed between 0 and 3" min="0" max="3"/>
169 </param>
170 <param name="bowtie_e" type="integer" value="99999999" label="Maximum sum of quality scores at mismatched positions in read alignments. This is also the value for the Bowtie -e option">
171 </param>
172 <param name="bowtie_m" type="integer" value="200" label="Discard alignments for reads with number of alignments greater than">
173 </param>
174 </when>
175 </conditional>
176 </macro>
177 <macro name="sampling_for_bam">
178 <param name="sampling_for_bam" type="boolean" truevalue="--sampling-for-bam" falsevalue="" checked="false" label="Use sampling for BAM">
179 <help> When RSEM generates a BAM file, instead of outputing all alignments a read has with their posterior probabilities, one alignment is sampled according to the posterior probabilities. The sampling procedure includes the alignment to the "noise" transcript, which does not appear in the BAM file. Only the sampled alignment has a weight of 1. All other alignments have weight 0. If the "noise" transcript is sampled, all alignments appeared in the BAM file should have weight 0. (Default: off)
180 </help>
181 </param>
182 </macro>
183 </macros>
184
185 <inputs>
186 <param name="sample" type="text" value="rsem_sample" label="Sample name" />
187 <conditional name="reference">
188 <param name="refSrc" type="select" label="RSEM Reference Source">
189 <option value="cached">Locally cached</option>
190 <option value="history">From your history</option>
191 </param>
192 <when value="cached">
193 <param name="index" type="select" label="Select RSEM reference" help="Select from a list of pre-indexed references. If you don't see anything consult the wrapper's documentation on how to create or download a reference">
194 <options from_data_table="rsem_indexes">
195 <filter type="sort_by" column="2" />
196 <validator type="no_options" message="No indexes are available" />
197 </options>
198 </param>
199 </when>
200 <when value="history">
201 <param name="rsem_ref" type="data" format="rsem_ref" label="RSEM reference" />
202 </when>
203 </conditional>
204 <conditional name="input">
205 <param name="format" type="select" label="RSEM Input file type">
206 <option value="fastq">FASTQ</option>
207 <option value="fasta">FASTA</option>
208 <option value="sam">SAM/BAM</option>
209 </param>
210 <when value="fastq">
211 <param name="fastq_select" size="15" type="select" label="FASTQ type" >
212 <option value="--phred33-quals" selected="true">phred33 qualities (default for sanger)</option>
213 <option value="--solexa-quals">solexa qualities</option>
214 <option value="--phred64-quals">phred64 qualities</option>
215 </param>
216 <conditional name="fastq">
217 <param name="matepair" type="select" label="Library type">
218 <option value="single">Single End Reads</option>
219 <option value="paired">Paired End Reads</option>
220 </param>
221 <when value="single">
222 <param name="singlefastq" type="data" format="fastq" label="FASTQ file" />
223 </when>
224 <when value="paired">
225 <param name="fastq1" type="data" format="fastq" label="Read 1 fastq file" />
226 <param name="fastq2" type="data" format="fastq" label="Read 2 fastq file" />
227 </when>
228 </conditional>
229 <expand macro="bowtie_options"/>
230 </when>
231 <when value="fasta">
232 <conditional name="fasta">
233 <param name="matepair" type="select" label="Library Type">
234 <option value="single">Single End Reads</option>
235 <option value="paired">Paired End Reads</option>
236 </param>
237 <when value="single">
238 <param name="singlefasta" type="data" format="fasta" label="fasta file" />
239 </when>
240 <when value="paired">
241 <param name="fasta1" type="data" format="fasta" label="Read 1 fasta file" />
242 <param name="fasta2" type="data" format="fasta" label="Read 2 fasta file" />
243 </when>
244 </conditional>
245 <expand macro="bowtie_options"/>
246 </when>
247 <when value="sam">
248 <!-- convert-sam-for-rsem /ref/mouse_125 input.sam -o input_for_rsem.sam -->
249 <param name="matepair" type="select" label="Library Type">
250 <option value="single">Single End Reads</option>
251 <option value="paired">Paired End Reads</option>
252 </param>
253 <param name="rsem_sam" type="data" format="rsem_sam" label="RSEM formatted SAM file" />
254 </when>
255 </conditional>
256 <expand macro="rsem_options"/>
257 <conditional name="rsem_outputs">
258 <param name="result_bams" type="select" label="Create bam results files"
259 help="In addition to the transcript-coordinate-based BAM file output, also output a BAM file with the read alignments in genomic coordinates" >
260 <option value="none">No BAM results files</option>
261 <option value="default" selected="true">Transcript BAM results file</option>
262 <option value="both">Transcript and genome BAM results files</option>
263 </param>
264 <when value="none"/>
265 <when value="default">
266 <expand macro="sampling_for_bam"/>
267 </when>
268 <when value="both">
269 <expand macro="sampling_for_bam"/>
270 </when>
271 </conditional>
272 </inputs>
273 <stdio>
274 <exit_code range="1:" level="fatal" description="Error Running RSEM" />
275 </stdio>
276 <outputs>
277 <data format="tabular" name="gene_abundances" label="${sample}.gene_abundances" from_work_dir="rsem_output.genes.results"/>
278 <data format="tabular" name="isoform_abundances" label="${sample}.isoform_abundances" from_work_dir="rsem_output.isoforms.results"/>
279 <data format="bam" name="transcript_bam" label="${sample}.transcript.bam" from_work_dir="rsem_output.transcript.bam" >
280 <filter>rsem_outputs['result_bams'] != "none"</filter>
281 </data>
282 <data format="bam" name="transcript_sorted_bam" label="${sample}.transcript.bam" from_work_dir="rsem_output.transcript.sorted.bam" >
283 <filter>rsem_outputs['result_bams'] != "none"</filter>
284 </data>
285 <data format="bam" name="genome_bam" label="${sample}.genome.bam" from_work_dir="rsem_output.genome.bam">
286 <filter>rsem_outputs['result_bams'] == "both"</filter>
287 </data>
288 <data format="bam" name="genome_sorted_bam" label="${sample}.genome.sorted.bam" from_work_dir="rsem_output.genome.sorted.bam">
289 <filter>rsem_outputs['result_bams'] == "both"</filter>
290 </data>
291 <data format="txt" name="log" label="${sample}.rsem_log"/>
292 </outputs>
293 <tests>
294 <test>
295 <param name="sample" value="rsem_sample"/>
296 <param name="refSrc" value="history"/>
297 <param name="rsem_ref" value="RSEM_ref_reference.zip" ftype="rsem_ref"/>
298 <param name="format" value="fastq"/>
299 <param name="matepair" value="single"/>
300 <param name="singlefastq" value="test.fastq" ftype="fastqsanger"/>
301 <param name="result_bams" value="none"/>
302 <output name="gene_abundances">
303 <assert_contents>
304 <has_text text="ENST00000423562,ENST00000438504,ENST00000488147,ENST00000538476,ENST00000541675" />
305 </assert_contents>
306 </output>
307 <output name="isoform_abundances">
308 <assert_contents>
309 <has_text text="ENST00000332831" />
310 </assert_contents>
311 </output>
312 <output name="log">
313 <assert_contents>
314 <has_text text="Expression Results are written" />
315 </assert_contents>
316 </output>
317 </test>
318 </tests>
319 <help>
320
321
322 RSEM HOME PAGE - http://deweylab.biostat.wisc.edu/rsem/
323
324 NAME
325 rsem-calculate-expression
326
327 SYNOPSIS
328 rsem-calculate-expression [options] upstream_read_file(s) reference_name sample_name
329 rsem-calculate-expression [options] --paired-end upstream_read_file/s downstream_read_file/s reference_name sample_name
330 rsem-calculate-expression [options] --sam/--bam [--paired-end] input reference_name sample_name
331
332 ARGUMENTS
333 upstream_read_files/s
334 Comma-separated list of files containing single-end reads or
335 upstream reads for paired-end data. By default, these files are
336 assumed to be in FASTQ format. If the --no-qualities option is
337 specified, then FASTA format is expected.
338
339 downstream_read_file/s
340 Comma-separated list of files containing downstream reads which are
341 paired with the upstream reads. By default, these files are assumed
342 to be in FASTQ format. If the --no-qualities option is specified,
343 then FASTA format is expected.
344
345 input
346 SAM/BAM formatted input file. If "-" is specified for the filename,
347 SAM/BAM input is instead assumed to come from standard input. RSEM
348 requires all alignments of the same read group together. For
349 paired-end reads, RSEM also requires the two mates of any alignment
350 be adjacent. See Description section for how to make input file obey
351 RSEM's requirements.
352
353 reference_name
354 The name of the reference used. The user must have run
355 'rsem-prepare-reference' with this reference_name before running
356 this program.
357
358 sample_name
359 The name of the sample analyzed. All output files are prefixed by
360 this name (e.g., sample_name.genes.results)
361
362 OPTIONS
363
364 --paired-end
365 Input reads are paired-end reads. (Default: off)
366
367 --no-qualities
368 Input reads do not contain quality scores. (Default: off)
369
370 --strand-specific
371 The RNA-Seq protocol used to generate the reads is strand specific,
372 i.e., all (upstream) reads are derived from the forward strand. This
373 option is equivalent to --forward-prob=1.0. With this option set, if
374 RSEM runs the Bowtie aligner, the '--norc' Bowtie option will be
375 used, which disables alignment to the reverse strand of transcripts.
376 (Default: off)
377
378 --sam
379 Input file is in SAM format. (Default: off)
380
381 --bam
382 Input file is in BAM format. (Default: off)
383
384 --sam-header-info [file]
385 RSEM reads header information from input by default. If this option
386 is on, header information is read from the specified file. For the
387 format of the file, please see SAM official website. (Default: "")
388
389 -p/--num-threads [int]
390 Number of threads to use. Both Bowtie and expression estimation will
391 use this many threads. (Default: 1)
392
393 --no-bam-output
394 Do not output any BAM file. (Default: off)
395
396 --output-genome-bam
397 Generate a BAM file, 'sample_name.genome.bam', with alignments
398 mapped to genomic coordinates and annotated with their posterior
399 probabilities. In addition, RSEM will call samtools (included in
400 RSEM package) to sort and index the bam file.
401 'sample_name.genome.sorted.bam' and
402 'sample_name.genome.sorted.bam.bai' will be generated. (Default:
403 off)
404
405 --sampling-for-bam
406 When RSEM generates a BAM file, instead of outputing all alignments
407 a read has with their posterior probabilities, one alignment is
408 sampled and outputed according to the posterior probabilities. If
409 the sampling result is that the read comes from the "noise"
410 transcript, nothing is outputed. (Default: off)
411
412 --calc-ci
413 Calculate 95% credibility intervals and posterior mean estimates.
414 (Default: off)
415
416 --seed-length [int]
417 Seed length used by the read aligner. Providing the correct value is
418 important for RSEM. If RSEM runs Bowtie, it uses this value for
419 Bowtie's seed length parameter. Any read with its or at least one of
420 its mates' (for paired-end reads) length less than this value will
421 be ignored. If the references are not added poly(A) tails, the
422 minimum allowed value is 5, otherwise, the minimum allowed value is
423 25. Note that this script will only check if the value less or equal than
424 5 and give a warning message if the value less than 25 but greter or equal than
425 5. (Default: 25)
426
427 --tag [string]
428 The name of the optional field used in the SAM input for identifying
429 a read with too many valid alignments. The field should have the
430 format [tagName]:i:[value], where a [value] bigger than 0 indicates
431 a read with too many alignments. (Default: "")
432
433 --bowtie-path [path]
434 The path to the bowtie executables. (Default: the path to the bowtie
435 executables is assumed to be in the user's PATH environment
436 variable)
437
438 --bowtie-n [int]
439 (Bowtie parameter) max # of mismatches in the seed. (Range: 0-3,
440 Default: 2)
441
442 --bowtie-e [int]
443 (Bowtie parameter) max sum of mismatch quality scores across the
444 alignment. (Default: 99999999)
445
446 --bowtie-m [int]
447 (Bowtie parameter) suppress all alignments for a read if greater then [int]
448 valid alignments exist. (Default: 200)
449
450 --bowtie-chunkmbs [int]
451 (Bowtie parameter) memory allocated for best first alignment
452 calculation (Default: 0 - use bowtie's default)
453
454 --phred33-quals
455 Input quality scores are encoded as Phred+33. (Default: on)
456
457 --phred64-quals
458 Input quality scores are encoded as Phred+64 (default for GA
459 Pipeline ver. less than 1.3). (Default: off)
460
461 --solexa-quals
462 Input quality scores are solexa encoded (from GA Pipeline ver. less
463 than 1.3). (Default: off)
464
465 --forward-prob [double]
466 Probability of generating a read from the forward strand of a
467 transcript. Set to 1 for a strand-specific protocol where all
468 (upstream) reads are derived from the forward strand, 0 for a
469 strand-specific protocol where all (upstream) read are derived from
470 the reverse strand, or 0.5 for a non-strand-specific protocol.
471 (Default: 0.5)
472
473 --fragment-length-min [int]
474 Minimum read/insert length allowed. This is also the value for the
475 bowtie -I option. (Default: 1)
476
477 --fragment-length-max [int]
478 Maximum read/insert length allowed. This is also the value for the
479 bowtie -X option. (Default: 1000)
480
481 --fragment-length-mean [double]
482 (single-end data only) The mean of the fragment length distribution,
483 which is assumed to be a Gaussian. (Default: -1, which disables use
484 of the fragment length distribution)
485
486 --fragment-length-sd [double]
487 (single-end data only) The standard deviation of the fragment length
488 distribution, which is assumed to be a Gaussian. (Default: 0, which
489 assumes that all fragments are of the same length, given by the
490 rounded value of --fragment-length-mean)
491
492 --estimate-rspd
493 Set this option if you want to estimate the read start position
494 distribution (RSPD) from data. Otherwise, RSEM will use a uniform
495 RSPD. (Default: off)
496
497 --num-rspd-bins [int]
498 Number of bins in the RSPD. Only relevant when '--estimate-rspd' is
499 specified. Use of the default setting is recommended. (Default: 20)
500
501 --ci-memory [int]
502 Maximum size (in memory, MB) of the auxiliary buffer used for
503 computing credibility intervals (CI). Set it larger for a faster CI
504 calculation. However, leaving 2 GB memory free for other usage is
505 recommended. (Default: 1024)
506
507 --keep-intermediate-files
508 Keep temporary files generated by RSEM. RSEM creates a temporary
509 directory, 'sample_name.temp', into which it puts all intermediate
510 output files. If this directory already exists, RSEM overwrites all
511 files generated by previous RSEM runs inside of it. By default,
512 after RSEM finishes, the temporary directory is deleted. Set this
513 option to prevent the deletion of this directory and the
514 intermediate files inside of it. (Default: off)
515
516 --time
517 Output time consumed by each step of RSEM to 'sample_name.time'.
518 (Default: off)
519
520 -q/--quiet
521 Suppress the output of logging information. (Default: off)
522
523 -h/--help
524 Show help information.
525
526 DESCRIPTION
527 In its default mode, this program aligns input reads against a reference
528 transcriptome with Bowtie and calculates expression values using the
529 alignments. RSEM assumes the data are single-end reads with quality
530 scores, unless the '--paired-end' or '--no-qualities' options are
531 specified. Users may use an alternative aligner by specifying one of the
532 --sam and --bam options, and providing an alignment file in the
533 specified format. However, users should make sure that they align
534 against the indices generated by 'rsem-prepare-reference' and the
535 alignment file satisfies the requirements mentioned in ARGUMENTS
536 section.
537
538 One simple way to make the alignment file satisfying RSEM's requirements
539 (assuming the aligner used put mates in a paired-end read adjacent) is
540 to use 'convert-sam-for-rsem' script. This script only accept SAM format
541 files as input. If a BAM format file is obtained, please use samtools to
542 convert it to a SAM file first. For example, if '/ref/mouse_125' is the
543 'reference_name' and the SAM file is named 'input.sam', you can run the
544 following command:
545
546 convert-sam-for-rsem /ref/mouse_125 input.sam -o input_for_rsem.sam
547
548 For details, please refer to 'convert-sam-for-rsem's documentation page.
549
550 The SAM/BAM format RSEM uses is v1.4. However, it is compatible with old
551 SAM/BAM format. However, RSEM cannot recognize 0x100 in the FLAG field.
552 In addition, RSEM requires SEQ and QUAL are not '*'.
553
554 The user must run 'rsem-prepare-reference' with the appropriate
555 reference before using this program.
556
557 For single-end data, it is strongly recommended that the user provide
558 the fragment length distribution parameters (--fragment-length-mean and
559 --fragment-length-sd). For paired-end data, RSEM will automatically
560 learn a fragment length distribution from the data.
561
562 Please note that some of the default values for the Bowtie parameters
563 are not the same as those defined for Bowtie itself.
564
565 The temporary directory and all intermediate files will be removed when
566 RSEM finishes unless '--keep-intermediate-files' is specified.
567
568 With the '--calc-ci' option, 95% credibility intervals and posterior
569 mean estimates will be calculated in addition to maximum likelihood
570 estimates.
571
572 OUTPUT
573 sample_name.genes.results
574 File containing gene level expression estimates. The format of each
575 line in this file is:
576
577 gene_id expected_counts tau_value [pmc_value tau_pme_value
578 tau_ci_lower_bound tau_ci_upper_bound] transcript_id_list
579
580 Fields are separated by the tab character. Fields within "[]" are
581 only presented if '--calc-ci' is set. pme stands for posterior mean
582 estimation. pmc stands for posterior mean counts. ci_lower_bound(l)
583 means the lower bound of the credibility intervals,
584 ci_upper_bound(u) means the upper bound of the credibility
585 intervals. So the credibility interval is [l, u].
586 'transcript_id_list' is a space-separated list of transcript_ids
587 belonging to the gene. If no gene information is provided, this file
588 has the same content as 'sample_name.isoforms.results'.
589
590 sample_name.isoforms.results
591 File containing isoform level expression values. The format of each
592 line in this file is:
593
594 transcript_id expected_counts tau_value [pmc_value tau_pme_value
595 tau_ci_lower_bound tau_ci_upper_bound] gene_id
596
597 Fields are separated by the tab character. 'gene_id' is the gene_id
598 of the gene which this transcript belongs to. If no gene information
599 is provided, 'gene_id' and 'transcript_id' are the same.
600
601 sample_name.transcript.bam, sample_name.transcript.sorted.bam and
602 sample_name.transcript.sorted.bam.bai
603 Only generated when --no-bam-output is not specified.
604
605 'sample_name.transcript.bam' is a BAM-formatted file of read
606 alignments in transcript coordinates. The MAPQ field of each
607 alignment is set to min(100, floor(-10 * log10(1.0 - w) + 0.5)),
608 where w is the posterior probability of that alignment being the
609 true mapping of a read. In addition, RSEM pads a new tag ZW:f:value,
610 where value is a single precision floating number representing the
611 posterior probability.
612
613 'sample_name.transcript.sorted.bam' and
614 'sample_name.transcript.sorted.bam.bai' are the sorted BAM file and
615 indices generated by samtools (included in RSEM package).
616
617 sample_name.genome.bam, sample_name.genome.sorted.bam and
618 sample_name.genome.sorted.bam.bai
619 Only generated when --no-bam-output is not specified and
620 --output-genome-bam is specified.
621
622 'sample_name.genome.bam' is a BAM-formatted file of read alignments
623 in genomic coordinates. Alignments of reads that have identical
624 genomic coordinates (i.e., alignments to different isoforms that
625 share the same genomic region) are collapsed into one alignment. The
626 MAPQ field of each alignment is set to min(100, floor(-10 *
627 log10(1.0 - w) + 0.5)), where w is the posterior probability of that
628 alignment being the true mapping of a read. In addition, RSEM pads a
629 new tag ZW:f:value, where value is a single precision floating
630 number representing the posterior probability. If an alignment is
631 spliced, a XS:A:value tag is also added, where value is either '+'
632 or '-' indicating the strand of the transcript it aligns to.
633
634 'sample_name.genome.sorted.bam' and
635 'sample_name.genome.sorted.bam.bai' are the sorted BAM file and
636 indices generated by samtools (included in RSEM package).
637
638 sample_name.sam.gz
639 Only generated when the input files are raw reads instead of SAM/BAM
640 format files
641
642 It is the gzipped SAM output produced by bowtie aligner.
643
644 sample_name.time
645 Only generated when --time is specified.
646
647 It contains time (in seconds) consumed by aligning reads, estimating
648 expression levels and calculating credibility intervals.
649
650 sample_name.stat
651 This is a folder instead of a file. All model related statistics are
652 stored in this folder. Use 'rsem-plot-model' can generate plots
653 using this folder.
654
655 EXAMPLES
656 Assume the path to the bowtie executables is in the user's PATH
657 environment variable. Reference files are under '/ref' with name
658 'mouse_125'.
659
660 1) '/data/mmliver.fq', single-end reads with quality scores. Quality
661 scores are encoded as for 'GA pipeline version >= 1.3'. We want to use 8
662 threads and generate a genome BAM file:
663
664 rsem-calculate-expression --phred64-quals \
665 -p 8 \
666 --output-genome-bam \
667 /data/mmliver.fq \
668 /ref/mouse_125 \
669 mmliver_single_quals
670
671 2) '/data/mmliver_1.fq' and '/data/mmliver_2.fq', paired-end reads with
672 quality scores. Quality scores are in SANGER format. We want to use 8
673 threads and do not generate a genome BAM file:
674
675 rsem-calculate-expression -p 8 \
676 --paired-end \
677 /data/mmliver_1.fq \
678 /data/mmliver_2.fq \
679 /ref/mouse_125 \
680 mmliver_paired_end_quals
681
682 3) '/data/mmliver.fa', single-end reads without quality scores. We want
683 to use 8 threads:
684
685 rsem-calculate-expression -p 8 \
686 --no-qualities \
687 /data/mmliver.fa \
688 /ref/mouse_125 \
689 mmliver_single_without_quals
690
691 4) Data are the same as 1). We want to take a fragment length
692 distribution into consideration. We set the fragment length mean to 150
693 and the standard deviation to 35. In addition to a BAM file, we also
694 want to generate credibility intervals. We allow RSEM to use 1GB of
695 memory for CI calculation:
696
697 rsem-calculate-expression --bowtie-path /sw/bowtie \
698 --phred64-quals \
699 --fragment-length-mean 150.0 \
700 --fragment-length-sd 35.0 \
701 -p 8 \
702 --output-genome-bam \
703 --calc-ci \
704 --ci-memory 1024 \
705 /data/mmliver.fq \
706 /ref/mouse_125 \
707 mmliver_single_quals
708
709 5) '/data/mmliver_paired_end_quals.bam', paired-end reads with quality
710 scores. We want to use 8 threads:
711
712 rsem-calculate-expression --paired-end \
713 --bam \
714 -p 8 \
715 /data/mmliver_paired_end_quals.bam \
716 /ref/mouse_125 \
717 mmliver_paired_end_quals
718 </help>
719 </tool>