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1 <tool name="SAM/BAM Hybrid Selection Metrics" id="PicardHsMetrics" version="1.56.0">
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2 <description>for targeted resequencing data</description>
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3 <command interpreter="python">
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4
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5 picard_wrapper.py -i "${input_file}" -d "${html_file.files_path}" -t "${html_file}" --datatype "${input_file.ext}"
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6 --baitbed "${bait_bed}" --targetbed "${target_bed}" -n "${out_prefix}" --tmpdir "${__new_file_path__}"
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7 -j "\$JAVA_JAR_PATH/CalculateHsMetrics.jar"
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8
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9 </command>
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10 <requirements><requirement type="package" version="1.56.0">picard</requirement></requirements>
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11 <inputs>
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12 <param format="sam,bam" name="input_file" type="data" label="SAM/BAM dataset to generate statistics for" />
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13 <param name="out_prefix" value="Picard HS Metrics" type="text" label="Title for the output file" help="Use to remind you what the job was for." size="80" />
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14 <param name="bait_bed" type="data" format="bed,interval" label="Bait intervals: Sequences for bait in the design" help="Note specific format requirements below!" size="80" />
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15 <param name="target_bed" type="data" format="bed,interval" label="Target intervals: Sequences for targets in the design" help="Note specific format requirements below!" size="80" />
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16 <!--
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17
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18 Users can be enabled to set Java heap size by uncommenting this option and adding '-x "$maxheap"' to the <command> tag.
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19 If commented out the heapsize defaults to the value specified within picard_wrapper.py
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20
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21 <param name="maxheap" type="select"
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22 help="If in doubt, try the default. If it fails with a complaint about java heap size, try increasing it please - larger jobs will require your own hardware."
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23 label="Java heap size">
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24 <option value="4G" selected = "true">4GB default </option>
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25 <option value="8G" >8GB use if 4GB fails</option>
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26 <option value="16G">16GB - try this if 8GB fails</option>
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27 </param>
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28
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29 -->
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30 </inputs>
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31 <outputs>
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32 <data format="html" name="html_file" label="${out_prefix}.html" />
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33 </outputs>
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34 <tests>
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35 <test>
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36 <!-- Uncomment this if maxheap parameter is enabled
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37 <param name="maxheap" value="8G" />
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38 -->
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39 <param name="out_prefix" value="HSMetrics" />
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40 <param name="input_file" value="picard_input_summary_alignment_stats.sam" ftype="sam" />
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41 <param name="bait_bed" value="picard_input_bait.bed" />
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42 <param name="target_bed" value="picard_input_bait.bed" />
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43 <output name="html_file" file="picard_output_hs_transposed_summary_alignment_stats.html" ftype="html" lines_diff="212"/>
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44 </test>
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45 </tests>
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46 <help>
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47
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48 .. class:: infomark
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49
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50 **Summary**
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51
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52 Calculates a set of Hybrid Selection specific metrics from an aligned SAM or BAM file.
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53
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54 .. class:: warnmark
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55
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56 **WARNING about bait and target files**
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57
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58 Picard is very fussy about the bait and target file format. If these are not exactly right, it will fail with an error something like:
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59
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60 Exception in thread "main" net.sf.picard.PicardException: Invalid interval record contains 6 fields: chr1 45787123 45787316 CASO_22G_25063 1000 +
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61
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62 If you see an error like that from this tool, please do NOT report it to any of the Galaxy mailing lists as it is not a bug!
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63 It means you must reformat your bait and target files. Galaxy cannot do that for you automatically unfortunately.
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64
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65 The required definition is described in the documentation at http://www.broadinstitute.org/gsa/wiki/index.php/Built-in_command-line_arguments
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66 and the sample provided looks like this:
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67
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68 chr1 1104841 1104940 + target_1
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69 chr1 1105283 1105599 + target_2
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70 chr1 1105712 1105860 + target_3
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71 chr1 1105960 1106119 + target_4
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72
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73 So your bait and target files MUST have 5 columns with chr, start, end, strand and name tab delimited and in exactly that order.
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74 Note that the Picard mandated sam header described in the documentation linked above is automagically added by the tool in Galaxy.
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75
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76 .. class:: infomark
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77
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78 **Picard documentation**
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79
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80 This is a Galaxy wrapper for CalculateHsMetrics.jar, a part of the external package Picard-tools_.
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81
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82 .. _Picard-tools: http://www.google.com/search?q=picard+samtools
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83
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84 -----
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85
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86 .. class:: infomark
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87
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88 **Inputs, outputs, and parameters**
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89
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90 Picard documentation says (reformatted for Galaxy):
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91
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92 Calculates a set of Hybrid Selection specific metrics from an aligned SAM or BAM file.
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93
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94 .. csv-table::
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95 :header-rows: 1
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96
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97 "Option", "Description"
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98 "BAIT_INTERVALS=File","An interval list file that contains the locations of the baits used. Required."
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99 "TARGET_INTERVALS=File","An interval list file that contains the locations of the targets. Required."
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100 "INPUT=File","An aligned SAM or BAM file. Required."
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101 "OUTPUT=File","The output file to write the metrics to. Required. Cannot be used in conjuction with option(s) METRICS_FILE (M)"
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102 "METRICS_FILE=File","Legacy synonym for OUTPUT, should not be used. Required. Cannot be used in conjuction with option(s) OUTPUT (O)"
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103 "CREATE_MD5_FILE=Boolean","Whether to create an MD5 digest for any BAM files created. Default value: false"
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104
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105 HsMetrics
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106
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107 The set of metrics captured that are specific to a hybrid selection analysis.
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108
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109 Output Column Definitions::
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110
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111 1. BAIT_SET: The name of the bait set used in the hybrid selection.
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112 2. GENOME_SIZE: The number of bases in the reference genome used for alignment.
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113 3. BAIT_TERRITORY: The number of bases which have one or more baits on top of them.
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114 4. TARGET_TERRITORY: The unique number of target bases in the experiment where target is usually exons etc.
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115 5. BAIT_DESIGN_EFFICIENCY: Target terrirtoy / bait territory. 1 == perfectly efficient, 0.5 = half of baited bases are not target.
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116 6. TOTAL_READS: The total number of reads in the SAM or BAM file examine.
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117 7. PF_READS: The number of reads that pass the vendor's filter.
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118 8. PF_UNIQUE_READS: The number of PF reads that are not marked as duplicates.
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119 9. PCT_PF_READS: PF reads / total reads. The percent of reads passing filter.
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120 10. PCT_PF_UQ_READS: PF Unique Reads / Total Reads.
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121 11. PF_UQ_READS_ALIGNED: The number of PF unique reads that are aligned with mapping score > 0 to the reference genome.
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122 12. PCT_PF_UQ_READS_ALIGNED: PF Reads Aligned / PF Reads.
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123 13. PF_UQ_BASES_ALIGNED: The number of bases in the PF aligned reads that are mapped to a reference base. Accounts for clipping and gaps.
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124 14. ON_BAIT_BASES: The number of PF aligned bases that mapped to a baited region of the genome.
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125 15. NEAR_BAIT_BASES: The number of PF aligned bases that mapped to within a fixed interval of a baited region, but not on a baited region.
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126 16. OFF_BAIT_BASES: The number of PF aligned bases that mapped to neither on or near a bait.
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127 17. ON_TARGET_BASES: The number of PF aligned bases that mapped to a targetted region of the genome.
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128 18. PCT_SELECTED_BASES: On+Near Bait Bases / PF Bases Aligned.
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129 19. PCT_OFF_BAIT: The percentage of aligned PF bases that mapped neither on or near a bait.
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130 20. ON_BAIT_VS_SELECTED: The percentage of on+near bait bases that are on as opposed to near.
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131 21. MEAN_BAIT_COVERAGE: The mean coverage of all baits in the experiment.
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132 22. MEAN_TARGET_COVERAGE: The mean coverage of targets that recieved at least coverage depth = 2 at one base.
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133 23. PCT_USABLE_BASES_ON_BAIT: The number of aligned, de-duped, on-bait bases out of the PF bases available.
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134 24. PCT_USABLE_BASES_ON_TARGET: The number of aligned, de-duped, on-target bases out of the PF bases available.
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135 25. FOLD_ENRICHMENT: The fold by which the baited region has been amplified above genomic background.
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136 26. ZERO_CVG_TARGETS_PCT: The number of targets that did not reach coverage=2 over any base.
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137 27. FOLD_80_BASE_PENALTY: The fold over-coverage necessary to raise 80% of bases in "non-zero-cvg" targets to the mean coverage level in those targets.
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138 28. PCT_TARGET_BASES_2X: The percentage of ALL target bases acheiving 2X or greater coverage.
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139 29. PCT_TARGET_BASES_10X: The percentage of ALL target bases acheiving 10X or greater coverage.
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140 30. PCT_TARGET_BASES_20X: The percentage of ALL target bases acheiving 20X or greater coverage.
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141 31. PCT_TARGET_BASES_30X: The percentage of ALL target bases acheiving 30X or greater coverage.
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142 32. HS_LIBRARY_SIZE: The estimated number of unique molecules in the selected part of the library.
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143 33. HS_PENALTY_10X: The "hybrid selection penalty" incurred to get 80% of target bases to 10X. This metric should be interpreted as: if I have a design with 10 megabases of target, and want to get 10X coverage I need to sequence until PF_ALIGNED_BASES = 10^6 * 10 * HS_PENALTY_10X.
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144 34. HS_PENALTY_20X: The "hybrid selection penalty" incurred to get 80% of target bases to 20X. This metric should be interpreted as: if I have a design with 10 megabases of target, and want to get 20X coverage I need to sequence until PF_ALIGNED_BASES = 10^6 * 20 * HS_PENALTY_20X.
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145 35. HS_PENALTY_30X: The "hybrid selection penalty" incurred to get 80% of target bases to 10X. This metric should be interpreted as: if I have a design with 10 megabases of target, and want to get 30X coverage I need to sequence until PF_ALIGNED_BASES = 10^6 * 30 * HS_PENALTY_30X.
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146
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147 .. class:: warningmark
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148
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149 **Warning on SAM/BAM quality**
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150
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151 Many SAM/BAM files produced externally and uploaded to Galaxy do not fully conform to SAM/BAM specifications. Galaxy deals with this by using the **LENIENT**
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152 flag when it runs Picard, which allows reads to be discarded if they're empty or don't map. This appears to be the only way to deal with SAM/BAM that cannot be parsed.
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153
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154
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155 </help>
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156 </tool>
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