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1 <tool id="bismark_bowtie" name="Bismark" version="0.7.12.1">
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0
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2 <!-- Wrapper compatible with Bismark version 0.7.11 -->
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3 <description>bisulfite mapper (bowtie)</description>
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4 <!--<version_command>bismark version</version_command>-->
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5 <requirements>
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6 <requirement type="set_environment">SCRIPT_PATH</requirement>
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2
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7 <requirement type="package" version="0.1.19">samtools</requirement>
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0
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8 <requirement type="package" version="0.12.8">bowtie</requirement>
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9 </requirements>
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10 <parallelism method="basic"></parallelism>
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11 <command interpreter="python">
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12 bismark_wrapper.py
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13
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14 --bismark_path \$SCRIPT_PATH
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15
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16 ##
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17 ## Bismark Genome Preparation, if desired.
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18 ##
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19
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20 ## Handle reference file.
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21 #if $refGenomeSource.genomeSource == "history":
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22 --own-file=$refGenomeSource.ownFile
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23 #else:
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24 --indexes-path ${refGenomeSource.index.fields.path}
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25 #end if
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26
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27
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28 ##
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29 ## Input parameters
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30 ##
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31
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32
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33 #if $singlePaired.sPaired == "single":
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34 --single-paired $singlePaired.input_singles
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35
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36 #if $singlePaired.input_singles.ext == "fastqillumina":
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37 --phred64-quals
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38 --fastq
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39 #elif $singlePaired.input_singles.ext == "fastqsanger":
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40 --fastq
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41 #elif $singlePaired.input_singles.ext == "fasta":
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42 --fasta
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43 #end if
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44 #else:
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45 --mate-paired
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46 #set $mate1 = list()
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47 #set $mate2 = list()
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48 #for $mate_pair in $singlePaired.mate_list
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49 $mate1.append( str($mate_pair.input_mate1) )
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50 $mate2.append( str($mate_pair.input_mate2) )
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51 #end for
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52
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53 --mate1 #echo ','.join($mate1)
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54 --mate2 #echo ','.join($mate2)
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55
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56 #if $singlePaired.mate_list[0].input_mate1.ext == "fastqillumina":
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57 --phred64-quals
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58 --fastq
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59 #elif $singlePaired.mate_list[0].input_mate1.ext == "fastqsanger":
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60 --fastq
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61 #elif $singlePaired.mate_list[0].input_mate1.ext == "fasta":
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62 --fasta
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63 #end if
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64
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65 -I $singlePaired.minInsert
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66 -X $singlePaired.maxInsert
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67 #end if
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68
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69
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70 ## for now hardcode the value for the required memory per thread in --best mode
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71 --chunkmbs 512
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72
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73
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74 #if $params.settingsType == "custom":
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75
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76 ## default 20
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77 --seed-len $params.seed_len
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78 ## default 0
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79 --seed-mismatches $params.seed_mismatches
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80
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81 ## default 70
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82 ##--maqerr $params.maqerr
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83
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84 ## default unlimited
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85 #if $params.qupto != 0:
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86 --qupto $params.qupto
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87 #end if
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88 #if $params.skip_reads != 0:
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89 --skip-reads $params.skip_reads
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90 #end if
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91
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92 #if $params.bismark_stdout:
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93 --stdout $output_stdout
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94 #end if
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95
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96 #if $params.isReportOutput:
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97 --output-report-file $report_file
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98 #end if
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99
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100 #end if
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101
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102 ##
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103 ## Output parameters.
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104 ##
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105 --output $output
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106 ##$suppress_header
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107
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108 #if str( $singlePaired.sPaired ) == "single"
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109 #if $output_unmapped_reads_l
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110 --output-unmapped-reads $output_unmapped_reads_l
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111 #end if
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112 #if $output_suppressed_reads_l
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113 --output-suppressed-reads $output_suppressed_reads_l
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114 #end if
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115 #else
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116 #if $output_unmapped_reads_l and $output_unmapped_reads_r
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117 --output-unmapped-reads-l $output_unmapped_reads_l
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118 --output-unmapped-reads-r $output_unmapped_reads_r
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119 #end if
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120 #if $output_suppressed_reads_l and $output_suppressed_reads_l
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121 --output-suppressed-reads-l $output_suppressed_reads_l
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122 --output-suppressed-reads-r $output_suppressed_reads_r
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123 #end if
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124 #end if
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125
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126 </command>
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127 <inputs>
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128 <conditional name="refGenomeSource">
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129 <param name="genomeSource" type="select" label="Will you select a reference genome from your history or use a built-in index?" help="Built-ins were indexed using default options">
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130 <option value="indexed">Use a built-in index</option>
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131 <option value="history">Use one from the history</option>
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132 </param>
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133 <when value="indexed">
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134 <param name="index" type="select" label="Select a reference genome" help="If your genome of interest is not listed, contact your Galaxy admin.">
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135 <options from_data_table="bowtie_indexes">
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136 <filter type="sort_by" column="2"/>
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137 <validator type="no_options" message="No indexes are available for the selected input dataset"/>
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138 </options>
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139 </param>
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140 </when> <!-- build-in -->
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141 <when value="history">
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142 <param name="ownFile" type="data" format="fasta" metadata_name="dbkey" label="Select the reference genome" />
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143 </when> <!-- history -->
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144 </conditional> <!-- refGenomeSource -->
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145
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146 <!-- Input Parameters -->
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147 <conditional name="singlePaired">
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148 <param name="sPaired" type="select" label="Is this library mate-paired?">
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149 <option value="single">Single-end</option>
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150 <option value="paired">Paired-end</option>
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151 </param>
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152 <when value="single">
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153 <param name="input_singles" type="data" format="fastqsanger,fastqillumina,fastq,fasta" label="FASTQ/FASTA file" help="FASTQ or FASTA files." />
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154 </when>
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155 <when value="paired">
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156 <repeat name="mate_list" title="Paired End Pairs" min="1">
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157 <param name="input_mate1" type="data" format="fastqsanger,fastqillumina,fastq,fasta" label="Mate pair 1" help="FASTQ or FASTA files." />
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158 <param name="input_mate2" type="data" format="fastqsanger,fastqillumina,fastq,fasta" label="Mate pair 2" help="FASTQ or FASTA files." />
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159 </repeat>
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160 <param name="minInsert" type="integer" value="0" label="Minimum insert size for valid paired-end alignments" />
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161 <param name="maxInsert" type="integer" value="500" label="Maximum insert size for valid paired-end alignments" />
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162 </when>
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163 </conditional>
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164
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165
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166 <conditional name="params">
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167 <param name="settingsType" type="select" label="Bismark settings to use" help="You can use the default settings or set custom values for any of Bismark's parameters.">
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168 <option value="default">Use Defaults</option>
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169 <option value="custom">Full parameter list</option>
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170 </param>
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171 <when value="default" />
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172 <!-- Full/advanced params. -->
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173 <when value="custom">
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174 <!-- -n -->
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175 <param name="seed_mismatches" type="select" label="The maximum number of mismatches permitted in the 'seed'">
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176 <option value="0">0</option>
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177 <option value="1">1</option>
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178 <option value="2" selected="true">2</option>
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179 <option value="3">3</option>
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180 </param>
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181 <!-- -l -->
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182 <param name="seed_len" type="integer" value="28" label="The 'seed length'; The number of bases of the high quality end of the read to which the maximum number of mismatches applies" />
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183 <!--
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184 <param name="maqerr" type="integer" value="70" label="Maximum permitted total of quality values at all mismatched read positions throughout the entire alignment, not just in the 'seed'." />
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185 -->
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186 <param name="qupto" type="integer" value="0" label="Only aligns the first N reads or read pairs from the input" help="Default is 0 and means 'no-limit'." />
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187 <param name="skip_reads" type="integer" value="0" label="Skip (i.e. do not align) the first N reads or read pairs from the input" />
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188
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189 <param name="suppressed_read_file" type="boolean" truevalue="true" falsevalue="false" checked="false" label="Write ambiguous reads to an extra output file" help="Write all reads which produce more than one valid alignment with the same number of lowest mismatches or other reads that fail to align uniquely." />
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190 <param name="unmapped_read_file" type="boolean" truevalue="true" falsevalue="false" checked="false" label="Write all reads that could not be aligned to a file" />
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191 <!-- output Options -->
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192 <param name="bismark_stdout" type="boolean" truevalue="true" falsevalue="false" checked="false" label="Write the bismark output and summary information to an extra file" />
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193 <param name="isReportOutput" type="boolean" truevalue="true" falsevalue="false" checked="false" label="Offer all report files concatenated in one file" />
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194 <!--end output options -->
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195 </when> <!-- full -->
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196 </conditional> <!-- params -->
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197 <!--
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198 <param name="suppress_header" type="boolean" truevalue="..suppress-header" falsevalue="" checked="false" label="Suppress the header in the output SAM file" help="Bowtie produces SAM with several lines of header information by default." />
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199 -->
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200 </inputs>
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201 <outputs>
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202 <data format="txt" name="report_file" label="${tool.name} on ${on_string}: Report">
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203 <filter>
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204 ((
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205 params['settingsType'] == "custom" and
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206 params['isReportOutput'] is True
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207 ))
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208 </filter>
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209 </data>
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210 <data format="txt" name="output_stdout" label="${tool.name} on ${on_string}: Summary">
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211 <filter>
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212 ((
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213 params['settingsType'] == "custom" and
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214 params['bismark_stdout'] is True
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215 ))
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216 </filter>
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217 </data>
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218
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219 <data format="bam" name="output" label="${tool.name} on ${on_string}: mapped reads">
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220 <actions>
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221 <conditional name="refGenomeSource.genomeSource">
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222 <when value="indexed">
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223 <action type="metadata" name="dbkey">
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224 <option type="from_data_table" name="bowtie2_indexes" column="1" offset="0">
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225 <filter type="param_value" column="0" value="#" compare="startswith" keep="False"/>
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226 <filter type="param_value" ref="refGenomeSource.index" column="0"/>
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227 </option>
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228 </action>
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229 </when>
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230 <when value="history">
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231 <action type="metadata" name="dbkey">
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232 <option type="from_param" name="refGenomeSource.ownFile" param_attribute="dbkey" />
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233 </action>
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234 </when>
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235 </conditional>
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236 </actions>
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237 </data>
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238
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239 <data format="fastq" name="output_suppressed_reads_l" label="${tool.name} on ${on_string}: suppressed reads (L)">
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240 <filter>
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241 ((
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242 params['settingsType'] == "custom" and
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243 params['suppressed_read_file'] is True
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244 ))
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245 </filter>
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246 <actions>
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247 <conditional name="singlePaired.sPaired">
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248 <when value="single">
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249 <action type="format">
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250 <option type="from_param" name="singlePaired.input_singles" param_attribute="ext" />
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251 </action>
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252 </when>
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253 <when value="paired">
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254 <action type="format">
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255 <option type="from_param" name="singlePaired.mate_list[0].input_mate1" param_attribute="ext" />
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256 </action>
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257 </when>
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258 </conditional>
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259 </actions>
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260 </data>
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261
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262 <data format="fastq" name="output_suppressed_reads_r" label="${tool.name} on ${on_string}: suppressed reads (R)">
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263 <filter>singlePaired['sPaired'] == "paired"</filter>
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264 <filter>params['settingsType'] == "custom"</filter>
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265 <filter>params['supressed_read_file'] is True</filter>
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266 <actions>
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267 <conditional name="singlePaired.sPaired">
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268 <when value="single">
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269 <action type="format">
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270 <option type="from_param" name="singlePaired.input_singles" param_attribute="ext" />
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271 </action>
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272 </when>
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273 <when value="paired">
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274 <action type="format">
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275 <option type="from_param" name="singlePaired.mate_list[0].input_mate1" param_attribute="ext" />
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276 </action>
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277 </when>
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278 </conditional>
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279 </actions>
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280 </data>
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281
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282 <!-- Outout unmapped reads -->
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283 <data format="fastq" name="output_unmapped_reads_l" label="${tool.name} on ${on_string}: unmapped reads (L)">
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284 <filter>
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285 ((
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286 params['settingsType'] == "custom" and
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287 params['unmapped_read_file'] is True
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288 ))
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289 </filter>
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290 <actions>
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291 <conditional name="singlePaired.sPaired">
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292 <when value="single">
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293 <action type="format">
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294 <option type="from_param" name="singlePaired.input_singles" param_attribute="ext" />
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295 </action>
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296 </when>
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297 <when value="paired">
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298 <action type="format">
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299 <option type="from_param" name="singlePaired.mate_list[0].input_mate1" param_attribute="ext" />
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300 </action>
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301 </when>
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302 </conditional>
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303 </actions>
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304 </data>
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305 <data format="fastq" name="output_unmapped_reads_r" label="${tool.name} on ${on_string}: unmapped reads (R)">
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306 <filter>singlePaired['sPaired'] == "paired"</filter>
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307 <filter>params['settingsType'] == "custom"</filter>
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308 <filter>params['unmapped_read_file'] is True</filter>
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309 <actions>
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310 <conditional name="singlePaired.sPaired">
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311 <when value="single">
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312 <action type="format">
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313 <option type="from_param" name="singlePaired.input_singles" param_attribute="ext" />
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314 </action>
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315 </when>
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316 <when value="paired">
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317 <action type="format">
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318 <option type="from_param" name="singlePaired.mate_list[0].input_mate1" param_attribute="ext" />
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319 </action>
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320 </when>
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321 </conditional>
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322 </actions>
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323 </data>
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324
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325
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326 </outputs>
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327
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328 <tests>
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329 </tests>
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330
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331 <help>
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332
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333 **What it does**
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334
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335 Bismark_ is a bisulfite mapper and methylation caller. Bismark takes in FastA or FastQ files and aligns the
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336 reads to a specified bisulfite genome. Sequence reads are transformed into a bisulfite converted forward strand
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337 version (C->T conversion) or into a bisulfite treated reverse strand (G->A conversion of the forward strand).
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338 Each of these reads are then aligned to bisulfite treated forward strand index of a reference genome
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339 (C->T converted) and a bisulfite treated reverse strand index of the genome (G->A conversion of the
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340 forward strand, by doing this alignments will produce the same positions). These 4 instances of Bowtie (1 or 2)
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341 are run in parallel. The sequence file(s) are then read in again sequence by sequence to pull out the original
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342 sequence from the genome and determine if there were any protected C's present or not.
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343
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344 .. _Bismark: http://www.bioinformatics.babraham.ac.uk/projects/bismark/
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345
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346 As of version 0.7.0 Bismark will only run 2 alignment threads for OT and OB in parallel, the 4 strand mode can be
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347 re-enabled by using non_directional mode.
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348
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349 It is developed by Krueger F and Andrews SR. at the Babraham Institute. Krueger F, Andrews SR. (2011) Bismark: a flexible aligner and methylation caller for Bisulfite-Seq applications. Bioinformatics, 27, 1571-2.
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350
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351 ------
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352
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353 **Know what you are doing**
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354
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355 .. class:: warningmark
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356
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357 There is no such thing (yet) as an automated gearshift in short read mapping. It is all like stick-shift driving in San Francisco. In other words = running this tool with default parameters will probably not give you meaningful results. A way to deal with this is to **understand** the parameters by carefully reading the `documentation`__ and experimenting. Fortunately, Galaxy makes experimenting easy.
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358
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359 .. __: http://www.bioinformatics.babraham.ac.uk/projects/bismark/
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360
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361
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362 .. class:: warningmark
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363
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364 Make sure all your input reads are in the correct and same format. If thats not the case please adjust/convert the filetype with galaxy's build-in converters.
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365
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366 ------
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367
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368 **Input formats**
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369
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370 Bismark accepts files in either Sanger FASTQ format (galaxy type *fastqsanger*), Illumina FASTQ format (galaxy type *fastqillumina*) or FASTA format (galaxy type *fasta*). Use the FASTQ Groomer to prepare your files.
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371
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372 ------
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373
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374 **A Note on Built-in Reference Genomes**
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375
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376 The default variant for all genomes is "Full", defined as all primary chromosomes (or scaffolds/contigs) including mitochondrial plus associated unmapped, plasmid, and other segments. When only one version of a genome is available in this tool, it represents the default "Full" variant. Some genomes will have more than one variant available. The "Canonical Male" or sometimes simply "Canonical" variant contains the primary chromosomes for a genome. For example a human "Canonical" variant contains chr1-chr22, chrX, chrY, and chrM. The "Canonical Female" variant contains the primary chromosomes excluding chrY.
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377
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378 ------
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379
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380 The final output of Bismark is in SAM format by default.
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381
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382 **Outputs**
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383
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384 The output is in SAM format, and has the following columns::
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385
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386 Column Description
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387 -------- --------------------------------------------------------
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388 1 QNAME seq-ID
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389 2 FLAG this flag tries to take the strand a bisulfite read
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390 originated from into account
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391 (this is different from ordinary DNA alignment flags!)
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392 3 RNAME chromosome
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393 4 POS start position
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394 5 MAPQ always 255
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395 6 CIGAR extended CIGAR string
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396 7 MRNM Mate Reference sequence NaMe ('=' if same as RNAME)
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397 8 MPOS 1-based Mate POSition
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398 9 ISIZE Inferred insert SIZE
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399 10 SEQ query SEQuence on the same strand as the reference
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400 11 QUAL Phred33 scale
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401 12 NM-tag edit distance to the reference)
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402 13 XX-tag base-by-base mismatches to the reference.
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403 This does not include indels.
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404 14 XM-tag methylation call string
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405 15 XR-tag read conversion state for the alignment
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406 16 XG-tag genome conversion state for the alignment
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407
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408
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409 Each read of paired-end alignments is written out in a separate line in the above format.
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410
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411
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412 It looks like this (scroll sideways to see the entire example)::
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413
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414 QNAME FLAG RNAME POS MAPQ CIAGR MRNM MPOS ISIZE SEQ QUAL OPT
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415 HWI-EAS91_1_30788AAXX:1:1:1761:343 4 * 0 0 * * 0 0 AAAAAAANNAAAAAAAAAAAAAAAAAAAAAAAAAAACNNANNGAGTNGNNNNNNNGCTTCCCACAGNNCTGG hhhhhhh;;hhhhhhhhhhh^hOhhhhghhhfhhhgh;;h;;hhhh;h;;;;;;;hhhhhhghhhh;;Phhh
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416 HWI-EAS91_1_30788AAXX:1:1:1578:331 4 * 0 0 * * 0 0 GTATAGANNAATAAGAAAAAAAAAAATGAAGACTTTCNNANNTCTGNANNNNNNNTCTTTTTTCAGNNGTAG hhhhhhh;;hhhhhhhhhhhhhhhhhhhhhhhhhhhh;;h;;hhhh;h;;;;;;;hhhhhhhhhhh;;hhVh
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417
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418 -------
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419
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420 **Bismark settings**
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421
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422 All of the options have a default value. You can change any of them. If any Bismark function is missing please contact the tool author or your Galaxy admin.
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423
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424 ------
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425
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426 **Bismark parameter list**
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427
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428 This is an exhaustive list of Bismark options.
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429
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430 Input::
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431
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432 --singles A comma- or space-separated list of files containing the reads to be aligned (e.g.
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433 lane1.fq,lane2.fq lane3.fq). Reads may be a mix of different lengths. Bismark will
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434 produce one mapping result and one report file per input file.
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435
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436 -1 mates1 Comma-separated list of files containing the #1 mates (filename usually includes
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437 "_1"), e.g. flyA_1.fq,flyB_1.fq). Sequences specified with this option must
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438 correspond file-for-file and read-for-read with those specified in mates2.
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439 Reads may be a mix of different lengths. Bismark will produce one mapping result
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440 and one report file per paired-end input file pair.
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441
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442 -2 mates2 Comma-separated list of files containing the #2 mates (filename usually includes
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443 "_2"), e.g. flyA_1.fq,flyB_1.fq). Sequences specified with this option must
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444 correspond file-for-file and read-for-read with those specified in mates1.
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445 Reads may be a mix of different lengths.
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446
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447 -q/--fastq The query input files (specified as mate1,mate2 or singles are FASTQ
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448 files (usually having extension .fg or .fastq). This is the default. See also
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449 --solexa-quals.
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450
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451 -f/--fasta The query input files (specified as mate1,mate2 or singles are FASTA
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452 files (usually havin extension .fa, .mfa, .fna or similar). All quality values
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453 are assumed to be 40 on the Phred scale.
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454
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455 -s/--skip INT Skip (i.e. do not align) the first INT reads or read pairs from the input.
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456
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457 -u/--upto INT Only aligns the first INT reads or read pairs from the input. Default: no limit.
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458
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459 --phred33-quals FASTQ qualities are ASCII chars equal to the Phred quality plus 33. Default: on.
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460
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461 --phred64-quals FASTQ qualities are ASCII chars equal to the Phred quality plus 64. Default: off.
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462
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463 --solexa-quals Convert FASTQ qualities from solexa-scaled (which can be negative) to phred-scaled
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464 (which can't). The formula for conversion is:
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465 phred-qual = 10 * log(1 + 10 ** (solexa-qual/10.0)) / log(10). Used with -q. This
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466 is usually the right option for use with (unconverted) reads emitted by the GA
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467 Pipeline versions prior to 1.3. Works only for Bowtie 1. Default: off.
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468
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469 --solexa1.3-quals Same as --phred64-quals. This is usually the right option for use with (unconverted)
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470 reads emitted by GA Pipeline version 1.3 or later. Default: off.
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471
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472
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473 Alignment::
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474
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475 -n/--seedmms INT The maximum number of mismatches permitted in the "seed", i.e. the first L base pairs
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476 of the read (where L is set with -l/--seedlen). This may be 0, 1, 2 or 3 and the
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477 default is 1. This option is only available for Bowtie 1 (for Bowtie 2 see -N).
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478
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479 -l/--seedlen The "seed length"; i.e., the number of bases of the high quality end of the read to
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480 which the -n ceiling applies. The default is 28. Bowtie (and thus Bismark) is faster for
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481 larger values of -l. This option is only available for Bowtie 1 (for Bowtie 2 see -L).
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482
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483 -e/--maqerr INT Maximum permitted total of quality values at all mismatched read positions throughout
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484 the entire alignment, not just in the "seed". The default is 70. Like Maq, bowtie rounds
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485 quality values to the nearest 10 and saturates at 30. This value is not relevant for
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486 Bowtie 2.
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487
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488 --chunkmbs INT The number of megabytes of memory a given thread is given to store path descriptors in
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489 --best mode. Best-first search must keep track of many paths at once to ensure it is
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490 always extending the path with the lowest cumulative cost. Bowtie tries to minimize the
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491 memory impact of the descriptors, but they can still grow very large in some cases. If
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492 you receive an error message saying that chunk memory has been exhausted in --best mode,
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493 try adjusting this parameter up to dedicate more memory to the descriptors. This value
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494 is not relevant for Bowtie 2. Default: 512.
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495
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496 -I/--minins INT The minimum insert size for valid paired-end alignments. E.g. if -I 60 is specified and
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497 a paired-end alignment consists of two 20-bp alignments in the appropriate orientation
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498 with a 20-bp gap between them, that alignment is considered valid (as long as -X is also
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499 satisfied). A 19-bp gap would not be valid in that case. Default: 0.
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500
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501 -X/--maxins INT The maximum insert size for valid paired-end alignments. E.g. if -X 100 is specified and
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502 a paired-end alignment consists of two 20-bp alignments in the proper orientation with a
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503 60-bp gap between them, that alignment is considered valid (as long as -I is also satisfied).
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504 A 61-bp gap would not be valid in that case. Default: 500.
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505
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506
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507
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508 Output::
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509
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510 --non_directional The sequencing library was constructed in a non strand-specific manner, alignments to all four
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511 bisulfite strands will be reported. Default: OFF.
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512
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513 (The current Illumina protocol for BS-Seq is directional, in which case the strands complementary
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514 to the original strands are merely theoretical and should not exist in reality. Specifying directional
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515 alignments (which is the default) will only run 2 alignment threads to the original top (OT)
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516 or bottom (OB) strands in parallel and report these alignments. This is the recommended option
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517 for sprand-specific libraries).
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518
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519 --sam-no-hd Suppress SAM header lines (starting with @). This might be useful when very large input files are
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520 split up into several smaller files to run concurrently and the output files are to be merged.
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521
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522 --quiet Print nothing besides alignments.
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523
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524 --vanilla Performs bisulfite mapping with Bowtie 1 and prints the 'old' output (as in Bismark 0.5.X) instead
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525 of SAM format output.
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526
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527 -un/--unmapped Write all reads that could not be aligned to a file in the output directory. Written reads will
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528 appear as they did in the input, without any translation of quality values that may have
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529 taken place within Bowtie or Bismark. Paired-end reads will be written to two parallel files with _1
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530 and _2 inserted in their filenames, i.e. _unmapped_reads_1.txt and unmapped_reads_2.txt. Reads
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531 with more than one valid alignment with the same number of lowest mismatches (ambiguous mapping)
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532 are also written to _unmapped_reads.txt unless the option --ambiguous is specified as well.
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533
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534 --ambiguous Write all reads which produce more than one valid alignment with the same number of lowest
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535 mismatches or other reads that fail to align uniquely to a file in the output directory.
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536 Written reads will appear as they did in the input, without any of the translation of quality
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537 values that may have taken place within Bowtie or Bismark. Paired-end reads will be written to two
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538 parallel files with _1 and _2 inserted in theit filenames, i.e. _ambiguous_reads_1.txt and
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539 _ambiguous_reads_2.txt. These reads are not written to the file specified with --un.
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540
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541 -o/--output_dir DIR Write all output files into this directory. By default the output files will be written into
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542 the same folder as the input file(s). If the specified folder does not exist, Bismark will attempt
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543 to create it first. The path to the output folder can be either relative or absolute.
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544
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545 --temp_dir DIR Write temporary files to this directory instead of into the same directory as the input files. If
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546 the specified folder does not exist, Bismark will attempt to create it first. The path to the
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547 temporary folder can be either relative or absolute.
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548
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549 </help>
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550 </tool>
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