Mercurial > repos > pjbriggs > pal_finder
annotate pal_finder_wrapper.xml @ 9:52dbe2089d14 draft default tip
Version 0.02.04.8 (update fastq subsetting).
| author | pjbriggs |
|---|---|
| date | Wed, 04 Jul 2018 06:05:52 -0400 |
| parents | 4e625d3672ba |
| children |
| rev | line source |
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| 9 | 1 <tool id="microsat_pal_finder" name="pal_finder" version="0.02.04.8"> |
| 2 | 2 <description>Find microsatellite repeat elements from sequencing reads and design PCR primers to amplify them</description> |
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3 <macros> |
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4 <import>pal_finder_macros.xml</import> |
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5 </macros> |
| 2 | 6 <requirements> |
| 7 <requirement type="package" version="0.02.04">pal_finder</requirement> | |
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8 <requirement type="package" version="2.7">python</requirement> |
| 2 | 9 <requirement type="package" version="1.65">biopython</requirement> |
| 10 <requirement type="package" version="2.8.1">pandaseq</requirement> | |
| 11 </requirements> | |
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12 <command detect_errors="exit_code"><![CDATA[ |
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13 @CONDA_PAL_FINDER_SCRIPT_DIR@ && |
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14 @CONDA_PAL_FINDER_DATA_DIR@ && |
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15 bash $__tool_directory__/pal_finder_wrapper.sh |
| 0 | 16 #if str( $platform.platform_type ) == "illumina" |
| 2 | 17 #set $paired_input_type = $platform.paired_input_type_conditional.paired_input_type |
| 18 #if $paired_input_type == "pair_of_files" | |
| 19 "$platform.paired_input_type_conditional.input_fastq_r1" | |
| 20 "$platform.paired_input_type_conditional.input_fastq_r2" | |
| 21 #else | |
| 22 "$platform.paired_input_type_conditional.input_fastq_pair.forward" | |
| 23 "$platform.paired_input_type_conditional.input_fastq_pair.reverse" | |
| 24 #end if | |
| 0 | 25 #else |
| 2 | 26 --454 "$platform.input_fasta" |
| 0 | 27 #end if |
| 28 $output_microsat_summary $output_pal_summary | |
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29 #if $report_bad_primer_ranges |
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30 --bad_primer_ranges "$output_bad_primer_read_ids" |
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31 #end if |
| 0 | 32 #if $keep_config_file |
| 2 | 33 --output_config_file "$output_config_file" |
| 0 | 34 #end if |
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35 --primer-prefix "$primer_prefix" |
| 0 | 36 --2merMinReps $min_2mer_repeats |
| 37 --3merMinReps $min_3mer_repeats | |
| 38 --4merMinReps $min_4mer_repeats | |
| 39 --5merMinReps $min_5mer_repeats | |
| 40 --6merMinReps $min_6mer_repeats | |
| 41 #if str( $primer.primer_options ) == "custom" | |
| 42 --primer-opt-size $primer.primer_opt_size | |
| 43 --primer-min-size $primer.primer_min_size | |
| 44 --primer-max-size $primer.primer_max_size | |
| 45 --primer-min-gc $primer.primer_min_gc | |
| 46 --primer-max-gc $primer.primer_max_gc | |
| 47 --primer-gc-clamp $primer.primer_gc_clamp | |
| 48 --primer-max-end-gc $primer.primer_max_end_gc | |
| 49 --primer-min-tm $primer.primer_min_tm | |
| 50 --primer-max-tm $primer.primer_max_tm | |
| 51 --primer-opt-tm $primer.primer_opt_tm | |
| 52 --primer-pair-max-diff-tm $primer.primer_pair_max_diff_tm | |
| 53 #end if | |
| 54 #if str( $mispriming.mispriming_options ) == "custom" | |
| 55 --primer-mispriming-library $mispriming.mispriming_library | |
| 56 #end if | |
| 2 | 57 #if str( $platform.platform_type ) == "illumina" |
| 58 #if $platform.filters | |
| 59 #for $filter in str($platform.filters).split(',') | |
| 60 $filter | |
| 61 --filter_microsats "$output_filtered_microsats" | |
| 62 #end for | |
| 63 #end if | |
| 64 #if str( $platform.assembly ) == '-assembly' | |
| 65 $platform.assembly "$output_assembly" | |
| 66 #end if | |
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67 #set $use_all_reads = $platform.subset_conditional.use_all_reads |
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68 #if str( $use_all_reads ) != "yes" |
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69 --subset "$platform.subset_conditional.subset" |
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70 #end if |
| 2 | 71 #end if |
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72 ]]></command> |
| 0 | 73 <inputs> |
| 74 <param name="primer_prefix" type="text" value="test" size="25" label="Primer prefix" help="This prefix will be added to the beginning of all primer names" /> | |
| 75 <conditional name="platform"> | |
| 76 <param name="platform_type" type="select" label="Sequencing platform used to generate data" help="Currently pal_finder only handles Illumina paired-end reads and 454 single-end reads" > | |
| 77 <option value="illumina" selected="true">Illumina</option> | |
| 78 <option value="454">454</option> | |
| 79 </param> | |
| 80 <when value="illumina"> | |
| 2 | 81 <conditional name="paired_input_type_conditional"> |
| 82 <param name="paired_input_type" type="select" label="Input Type"> | |
| 83 <option value="pair_of_files" selected="true">Pair of datasets</option> | |
| 84 <option value="collection">Dataset collection pair</option> | |
| 85 </param> | |
| 86 <when value="pair_of_files"> | |
| 87 <param name="input_fastq_r1" type="data" format="fastqsanger" | |
| 88 label="Illumina fastq file (read 1)" /> | |
| 89 <param name="input_fastq_r2" type="data" format="fastqsanger" | |
| 90 label="Illumina fastq file (read 2)" /> | |
| 91 </when> | |
| 92 <when value="collection"> | |
| 93 <param name="input_fastq_pair" format="fastqsanger" | |
| 94 type="data_collection" collection_type="paired" | |
| 95 label="Select FASTQ dataset collection with R1/R2 pair" /> | |
| 96 </when> | |
| 97 </conditional> | |
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98 <conditional name="subset_conditional"> |
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99 <param name="use_all_reads" type="boolean" label="Use all reads for microsatellite detection?" checked="True" truevalue="yes" falsevalue="no" /> |
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100 <when value="no"> |
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101 <param name="subset" type="text" value="0.5" label="Number or fraction of reads to use" help="Either an integer number of reads or a decimal fraction (e.g. 0.5 to select 50% of reads)" /> |
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102 </when> |
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103 <when value="yes" /> |
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104 </conditional> |
| 2 | 105 <param name="filters" type="select" display="checkboxes" |
| 106 multiple="True" label="Filters to apply to the pal_finder results" | |
| 107 help="Apply none, one or more filters to refine results"> | |
| 108 <option value="-primers" selected="True">Only include loci with designed primers</option> | |
| 109 <option value="-occurrences" selected="True">Exclude loci where the primer sequences occur more than once in the reads</option> | |
| 110 <option value="-rankmotifs" selected="True">Only include loci with 'perfect' motifs, and rank by motif size</option> | |
| 111 </param> | |
| 112 <param name="assembly" type="boolean" | |
| 113 checked="True" truevalue="-assembly" falsevalue="" | |
| 114 label="Use PANDAseq to assemble paired-end reads and confirm primer sequences are present in high-quality assembly" /> | |
| 0 | 115 </when> |
| 116 <when value="454"> | |
| 117 <param name="input_fasta" type="data" format="fasta" label="454 fasta file with raw reads" /> | |
| 118 </when> | |
| 119 </conditional> | |
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120 <param name="min_2mer_repeats" type="integer" value="6" label="Minimum number of 2-mer repeat units to detect" min="1" help="Must detect at least one repeat of this n-mer unit" /> |
| 0 | 121 <param name="min_3mer_repeats" type="integer" value="0" label="Minimum number of 3-mer repeat units" help="Set to zero to ignore repeats of this n-mer unit" /> |
| 122 <param name="min_4mer_repeats" type="integer" value="0" label="Minimum number of 4-mer repeat units" help="Set to zero to ignore repeats of this n-mer unit" /> | |
| 123 <param name="min_5mer_repeats" type="integer" value="0" label="Minimum number of 5-mer repeat units" help="Set to zero to ignore repeats of this n-mer unit" /> | |
| 124 <param name="min_6mer_repeats" type="integer" value="0" label="Minimum number of 6-mer repeat units" help="Set to zero to ignore repeats of this n-mer unit" /> | |
| 125 <conditional name="mispriming"> | |
| 126 <param name="mispriming_options" type="select" label="Mispriming library to use" help="Specify file of nucleotide sequences to avoid amplifying (PRIMER_MISPRIMING_LIBRARY)"> | |
| 127 <option value="default">Default from pal_finder</option> | |
| 128 <option value="custom">Custom sequences from history</option> | |
| 129 </param> | |
| 130 <when value="default"> | |
| 131 </when> | |
| 132 <when value="custom"> | |
| 133 <param name="mispriming_library" type="data" format="fasta" label="Select mispriming library from history" help="Fasta file containing sequences to avoid amplifying" /> | |
| 134 </when> | |
| 135 </conditional> | |
| 136 <conditional name="primer"> | |
| 137 <param name="primer_options" type="select" label="Primer settings to use" help="Advanced users can customise the settings for primer3 for more control"> | |
| 138 <option value="default">Defaults for pal_finder</option> | |
| 139 <option value="custom">Custom</option> | |
| 140 </param> | |
| 141 <when value="custom"> | |
| 142 <param name="primer_opt_size" type="integer" value="20" | |
| 143 label="Optimum length (in bases) of a primer (PRIMER_OPT_SIZE)" | |
| 144 help="Primer3 will attempt to pick primers close to this length" /> | |
| 145 <param name="primer_min_size" type="integer" value="18" | |
| 146 label="Minimum acceptable length (in bases) of a primer (PRIMER_MIN_SIZE)" | |
| 147 help="Must be greater than 0 and less than or equal to PRIMER_MAX_SIZE" /> | |
| 148 <param name="primer_max_size" type="integer" value="30" | |
| 149 label="Maximum acceptable length (in bases) of a primer (PRIMER_MAX_SIZE)" | |
| 150 help="Currently this parameter cannot be larger than 35. This limit is governed by maximum oligo size for which primer3's melting-temperature is valid" /> | |
| 151 <param name="primer_min_gc" type="float" value="30.0" | |
| 152 label="Minimum allowable percentage of Gs and Cs in any primer (PRIMER_MIN_GC)" /> | |
| 153 <param name="primer_max_gc" type="float" value="80.0" | |
| 154 label="Maximum allowable percentage of Gs and Cs in any primer (PRIMER_MAX_GC)" /> | |
| 155 <param name="primer_gc_clamp" type="integer" value="2" | |
| 156 label="Specify number of consecutive Gs and Cs at 3' end of both the left and right primer (PRIMER_GC_CLAMP)" /> | |
| 157 <param name="primer_max_end_gc" type="integer" value="5" | |
| 158 label="Maximum number of Gs or Cs allowed in last five 3' bases of a left or right primer (PRIMER_MAX_END_GC)" /> | |
| 159 <param name="primer_min_tm" type="float" value="58.0" | |
| 160 label="Minimum acceptable melting temperature for a primer oligo (PRIMER_MIN_TM)" | |
| 161 help="Temperature should be in degrees Celsius" /> | |
| 162 <param name="primer_max_tm" type="float" value="65.0" | |
| 163 label="Maximum acceptable melting temperature for a primer oligo (PRIMER_MAX_TM)" | |
| 164 help="Temperature should be in degrees Celsius" /> | |
| 165 <param name="primer_opt_tm" type="float" value="62.0" | |
| 166 label="Optimum melting temperature for a primer (PRIMER_OPT_TM)" | |
| 167 help="Temperature should be in degrees Celsius" /> | |
| 168 <param name="primer_pair_max_diff_tm" type="float" value="2.0" | |
| 169 label="Maximum acceptable difference between melting temperatures of left and right primers (PRIMER_PAIR_MAX_DIFF_TM)" | |
| 170 help="Temperature should be in degrees Celsius" /> | |
| 171 </when> | |
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172 <when value="default" /> |
| 0 | 173 </conditional> |
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174 <param name="report_bad_primer_ranges" type="boolean" truevalue="True" falsevalue="False" label="Output IDs for input reads which generate bad primer product size ranges" help="Can be used to screen reads in input Fastqs " /> |
| 0 | 175 <param name="keep_config_file" type="boolean" truevalue="True" falsevalue="False" |
| 176 label="Output the config file to the history" | |
| 177 help="Can be used to run pal_finder outside of Galaxy" /> | |
| 178 </inputs> | |
| 179 <outputs> | |
| 2 | 180 <data name="output_pal_summary" format="tabular" label="${tool.name} on ${on_string} for ${primer_prefix}: all microsatellites (full details)" /> |
| 181 <data name="output_filtered_microsats" format="tabular" label="${tool.name} on ${on_string} for ${primer_prefix}: filtered microsatellites (full details)"> | |
| 182 <filter>platform['platform_type'] == 'illumina' and platform['filters'] is not None</filter> | |
| 0 | 183 </data> |
| 2 | 184 <data name="output_microsat_summary" format="txt" label="${tool.name} on ${on_string} for ${primer_prefix}: summary of microsatellite types" /> |
| 185 <data name="output_assembly" format="tabular" label="${tool.name} on ${on_string} for ${primer_prefix}: assembly"> | |
| 186 <filter>platform['assembly'] is True</filter> | |
| 187 </data> | |
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188 <data name="output_bad_primer_read_ids" format="tabular" label="${tool.name} on ${on_string} for ${primer_prefix}: read IDs generating bad primer ranges"> |
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189 <filter>report_bad_primer_ranges is True</filter> |
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190 </data> |
| 2 | 191 <data name="output_config_file" format="txt" label="${tool.name} on ${on_string} for ${primer_prefix}: config file"> |
| 0 | 192 <filter>keep_config_file is True</filter> |
| 193 </data> | |
| 194 </outputs> | |
| 195 <tests> | |
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196 <!-- Test with Illumina input --> |
| 0 | 197 <test> |
| 198 <param name="platform_type" value="illumina" /> | |
| 199 <param name="input_fastq_r1" value="illuminaPE_r1.fq" ftype="fastqsanger" /> | |
| 200 <param name="input_fastq_r2" value="illuminaPE_r2.fq" ftype="fastqsanger" /> | |
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201 <expand macro="output_illumina_microsat_summary" /> |
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202 <output name="output_pal_summary" compare="re_match" file="illuminaPE_microsats.out.re_match" /> |
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203 <output name="output_filtered_microsats" compare="re_match" file="illuminaPE_filtered_microsats.out.re_match" /> |
| 2 | 204 <output name="output_assembly" file="illuminaPE_assembly_after_filters.out" /> |
| 205 </test> | |
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206 <!-- Test with Illumina input as dataset pair --> |
| 2 | 207 <test> |
| 208 <param name="platform_type" value="illumina" /> | |
| 209 <param name="paired_input_type" value="collection" /> | |
| 210 <param name="input_fastq_pair"> | |
| 211 <collection type="paired"> | |
| 212 <element name="forward" value="illuminaPE_r1.fq" ftype="fastqsanger" /> | |
| 213 <element name="reverse" value="illuminaPE_r2.fq" ftype="fastqsanger" /> | |
| 214 </collection> | |
| 215 </param> | |
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216 <expand macro="output_illumina_microsat_summary" /> |
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217 <output name="output_pal_summary" compare="re_match" file="illuminaPE_microsats.out.re_match" /> |
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218 <output name="output_filtered_microsats" compare="re_match" file="illuminaPE_filtered_microsats.out.re_match" /> |
| 2 | 219 <output name="output_assembly" file="illuminaPE_assembly_after_filters.out" /> |
| 220 </test> | |
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221 <!-- Test with Illumina input filter to loci with PandaSEQ assembly |
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222 ('-assembly' option) |
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223 --> |
| 2 | 224 <test> |
| 225 <param name="platform_type" value="illumina" /> | |
| 226 <param name="filters" value="" /> | |
| 227 <param name="input_fastq_r1" value="illuminaPE_r1.fq" ftype="fastqsanger" /> | |
| 228 <param name="input_fastq_r2" value="illuminaPE_r2.fq" ftype="fastqsanger" /> | |
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229 <expand macro="output_illumina_microsat_summary" /> |
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230 <output name="output_pal_summary" compare="re_match" file="illuminaPE_microsats.out.re_match" /> |
| 2 | 231 <output name="output_assembly" file="illuminaPE_assembly.out" /> |
| 232 </test> | |
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233 <!-- Test with Illumina input filter to loci with primers |
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234 ('-primers' option) --> |
| 2 | 235 <test> |
| 236 <param name="platform_type" value="illumina" /> | |
| 237 <param name="filters" value="-primers" /> | |
| 238 <param name="assembly" value="false" /> | |
| 239 <param name="input_fastq_r1" value="illuminaPE_r1.fq" ftype="fastqsanger" /> | |
| 240 <param name="input_fastq_r2" value="illuminaPE_r2.fq" ftype="fastqsanger" /> | |
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241 <expand macro="output_illumina_microsat_summary" /> |
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242 <output name="output_pal_summary" compare="re_match" file="illuminaPE_microsats.out.re_match" /> |
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243 <output name="output_filtered_microsats" compare="re_match" file="illuminaPE_filtered_microsats_primers.out.re_match" /> |
| 2 | 244 </test> |
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245 <!-- Test with Illumina input filter to loci which appear only once |
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246 ('-occurrences' option) --> |
| 2 | 247 <test> |
| 248 <param name="platform_type" value="illumina" /> | |
| 249 <param name="filters" value="-occurrences" /> | |
| 250 <param name="assembly" value="false" /> | |
| 251 <param name="input_fastq_r1" value="illuminaPE_r1.fq" ftype="fastqsanger" /> | |
| 252 <param name="input_fastq_r2" value="illuminaPE_r2.fq" ftype="fastqsanger" /> | |
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253 <expand macro="output_illumina_microsat_summary" /> |
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254 <output name="output_pal_summary" compare="re_match" file="illuminaPE_microsats.out.re_match" /> |
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255 <output name="output_filtered_microsats" compare="re_match" file="illuminaPE_filtered_microsats_occurrences.out.re_match" /> |
| 2 | 256 </test> |
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257 <!-- Test with Illumina input filter and rank loci with perfect motifs |
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258 ('-rankmotifs' option) --> |
| 2 | 259 <test> |
| 260 <param name="platform_type" value="illumina" /> | |
| 261 <param name="filters" value="-rankmotifs" /> | |
| 262 <param name="assembly" value="false" /> | |
| 263 <param name="input_fastq_r1" value="illuminaPE_r1.fq" ftype="fastqsanger" /> | |
| 264 <param name="input_fastq_r2" value="illuminaPE_r2.fq" ftype="fastqsanger" /> | |
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265 <expand macro="output_illumina_microsat_summary" /> |
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266 <output name="output_pal_summary" compare="re_match" file="illuminaPE_microsats.out.re_match" /> |
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267 <output name="output_filtered_microsats" compare="re_match" file="illuminaPE_filtered_microsats_rankmotifs.out.re_match" /> |
| 0 | 268 </test> |
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269 <!-- Test with Illumina input using subset of reads --> |
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270 <test> |
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271 <param name="platform_type" value="illumina" /> |
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272 <param name="filters" value="" /> |
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273 <param name="assembly" value="false" /> |
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274 <param name="use_all_reads" value="no" /> |
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275 <param name="subset" value="0.5" /> |
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276 <param name="input_fastq_r1" value="illuminaPE_r1.fq" ftype="fastqsanger" /> |
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277 <param name="input_fastq_r2" value="illuminaPE_r2.fq" ftype="fastqsanger" /> |
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278 <expand macro="output_illumina_microsat_subset_summary" /> |
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279 <output name="output_pal_summary" compare="re_match" file="illuminaPE_microsats_subset.out.re_match" /> |
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280 </test> |
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281 <!-- Test with Illumina input filter that doesn't find any |
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282 microsatellites --> |
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283 <test expect_failure="true"> |
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284 <param name="platform_type" value="illumina" /> |
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285 <param name="filters" value="" /> |
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286 <param name="assembly" value="false" /> |
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287 <param name="min_2mer_repeats" value="8" /> |
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288 <param name="input_fastq_r1" value="illuminaPE_r1_no_microsats.fq" ftype="fastqsanger" /> |
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289 <param name="input_fastq_r2" value="illuminaPE_r2_no_microsats.fq" ftype="fastqsanger" /> |
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290 <assert_stderr> |
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291 <has_text text="pal_finder failed to locate any microsatellites" /> |
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292 </assert_stderr> |
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293 </test> |
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294 <!-- Test with Illumina input generating bad ranges --> |
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295 <test> |
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296 <param name="platform_type" value="illumina" /> |
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297 <param name="filters" value="" /> |
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298 <param name="assembly" value="false" /> |
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299 <param name="min_2mer_repeats" value="8" /> |
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300 <param name="input_fastq_r1" value="illuminaPE_r1_bad_ranges.fq" ftype="fastqsanger" /> |
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301 <param name="input_fastq_r2" value="illuminaPE_r2_bad_ranges.fq" ftype="fastqsanger" /> |
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302 <param name="min_2mer_repeats" value="8" /> |
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303 <param name="min_3mer_repeats" value="8" /> |
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304 <param name="min_4mer_repeats" value="8" /> |
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305 <param name="min_5mer_repeats" value="8" /> |
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306 <param name="min_6mer_repeats" value="8" /> |
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307 <param name="primer_options" value="custom" /> |
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308 <param name="primer_opt_size" value="25" /> |
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309 <param name="primer_min_size" value="21" /> |
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310 <param name="primer_max_size" value="30" /> |
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311 <param name="primer_min_gc" value="40.0" /> |
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312 <param name="primer_max_gc" value="60.0" /> |
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313 <param name="primer_gc_clamp" value="3" /> |
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314 <param name="primer_max_end_gc" value="5" /> |
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315 <param name="primer_min_tm" value="60.0" /> |
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316 <param name="primer_max_tm" value="80.0" /> |
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317 <param name="primer_opt_tm" value="68.0" /> |
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318 <param name="primer_pair_max_diff_tm" value="3.0" /> |
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319 <param name="report_bad_primer_ranges" value="true" /> |
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320 <expand macro="output_illumina_microsat_summary_bad_ranges" /> |
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321 <output name="output_pal_summary" compare="re_match" file="illuminaPE_microsats_bad_ranges.out.re_match" /> |
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322 <output name="output_bad_primer_read_ids" file="illuminaPE_bad_primer_read_ids.out" /> |
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323 </test> |
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324 <!-- Test with bad n-mers specified --> |
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325 <test expect_failure="true"> |
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326 <param name="platform_type" value="illumina" /> |
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327 <param name="filters" value="" /> |
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328 <param name="assembly" value="false" /> |
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329 <param name="min_2mer_repeats" value="8" /> |
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330 <param name="min_3mer_repeats" value="8" /> |
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331 <param name="min_4mer_repeats" value="0" /> |
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332 <param name="min_5mer_repeats" value="8" /> |
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333 <param name="min_6mer_repeats" value="8" /> |
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334 <param name="input_fastq_r1" value="illuminaPE_r1_no_microsats.fq" ftype="fastqsanger" /> |
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335 <param name="input_fastq_r2" value="illuminaPE_r2_no_microsats.fq" ftype="fastqsanger" /> |
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336 <assert_stderr> |
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337 <has_text text="Minimum number of 4-mers cannot be zero if number of 5-mers is non-zero" /> |
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338 </assert_stderr> |
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339 </test> |
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340 <!-- Test with 454 input --> |
| 0 | 341 <test> |
| 342 <param name="platform_type" value="454" /> | |
| 343 <param name="input_fasta" value="454_in.fa" ftype="fasta" /> | |
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344 <expand macro="output_454_microsat_summary" /> |
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345 <output name="output_pal_summary" compare="re_match" file="454_microsats.out.re_match" /> |
| 0 | 346 </test> |
| 347 </tests> | |
| 348 <help> | |
| 349 .. class:: infomark | |
| 350 | |
| 351 **What it does** | |
| 352 | |
| 353 This tool runs the pal_finder program, which finds microsatellite repeat elements | |
| 354 directly from raw 454 or Illumina paired-end sequencing reads. It then designs PCR | |
| 355 primers to amplify these repeat loci (Potentially Amplifiable Loci: PAL). | |
| 356 | |
| 2 | 357 Optionally for Illumina data, one or more filters can be applied to the output from |
| 358 pal_finder to: | |
| 359 | |
| 360 * Only include loci with designed primers | |
| 361 * Exclude loci where the primer sequences occur more than once in the reads | |
| 362 * Only include loci with 'perfect' motifs (and rank by motif size,largest to | |
| 363 smallest) | |
| 364 * Use PANDAseq to assemble paired-end reads and confirm primer sequences are | |
| 365 present in high-quality assembly | |
| 0 | 366 |
| 367 Pal_finder runs the primer3_core program; information on the settings used in | |
| 368 primer3_core can be found in the Primer3 manual at | |
| 369 http://primer3.sourceforge.net/primer3_manual.htm | |
| 370 | |
| 371 ------------- | |
| 372 | |
| 373 .. class:: infomark | |
| 374 | |
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375 **Known issues** |
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376 |
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377 .. class:: warning |
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378 |
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379 **Low number of reads used for microsatellite detection/bad primer product size ranges** |
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380 |
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381 For some datasets pal_finder may generate 'bad' product size ranges (where the |
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382 lower limit exceeds the upper limit) for one or more reads, for input into |
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383 primer3_core. In these cases primer3_core will terminate prematurely, which can |
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384 result in a substantially lower number of reads being used for microsatellite |
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385 detection and potentially sub-optimal primer design. |
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386 |
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387 The number of reads generating the bad size ranges are reported in the |
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388 *Summary of microsat types* output dataset as 'readsWithBadRanges'. Ideally |
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389 the reported value should be zero. |
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390 |
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391 The conditions which cause this issue within pal_finder are still unclear, |
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392 however we believe it to be associated with short or low quality reads. If this |
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393 problem affects your data then: |
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394 |
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395 * Ensure that the input data are sufficiently trimmed and filtered (using |
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396 e.g. the Trimmomatic tool) before rerunning pal_finder. |
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397 |
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398 * A list of read IDs for which pal_finder generates bad product size ranges can |
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399 be output by turning on *Output IDs for input reads which generate bad primer |
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400 ranges*. This outputs an additional dataset with a list of read IDs which can |
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401 be used to remove read pairs from the input Fastq files (using e.g. the *Filter |
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402 sequences by ID* tool) before rerunning pal_finder. |
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403 |
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404 .. class:: warning |
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405 |
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406 **Pal_finder takes a long time to run for large input datasets** |
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407 |
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408 pal_finder was originally developed using MiSeq data, and is not optimised for |
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409 working with the larger Fastqs that are output from other platforms such as |
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410 HiSeq and NextSeq. As a consequence pal_finder may take a very long time to |
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411 complete when operating on larger datasets. |
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412 |
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413 If this is a problem then the tool can be run using a subset of the input reads |
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414 by unchecking the *Use all reads...* option and entering either an integer number |
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415 of reads to use, or a decimal fraction (e.g. 0.5 will select 50% of the reads). |
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416 |
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417 ------------- |
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418 |
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419 .. class:: infomark |
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420 |
| 0 | 421 **Credits** |
| 422 | |
| 423 This Galaxy tool has been developed by Peter Briggs within the Bioinformatics Core | |
| 424 Facility at the University of Manchester. It runs the pal_finder package which can be | |
| 425 obtained from http://sourceforge.net/projects/palfinder/: | |
| 426 | |
| 427 * PLoS One. 2012; 7(2): e30953 "Rapid Microsatellite Identification from Illumina Paired-End | |
| 428 Genomic Sequencing in Two Birds and a Snake" Todd A. Castoe, Alexander W. Poole, A. P. | |
| 429 Jason de Koning, Kenneth L. Jones, Diana F. Tomback, Sara J. Oyler-McCance, Jennifer A. | |
| 430 Fike, Stacey L. Lance, Jeffrey W. Streicher, Eric N. Smith, and David D. Pollock | |
| 431 | |
| 432 The paper is available at http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3279355/ | |
| 433 | |
| 434 This tool is compatible with pal_finder version 0.02.04, which in turn runs the | |
| 435 primer3_core program (version 2.0.0-alpha is required, available from | |
| 436 http://primer3.sourceforge.net/releases.php): | |
| 437 | |
| 438 * Steve Rozen and Helen J. Skaletsky (2000) "Primer3 on the WWW for general users and for | |
| 439 biologist programmers". In: Krawetz S, Misener S (eds) Bioinformatics Methods and | |
| 440 Protocols: Methods in Molecular Biology. Humana Press, Totowa, NJ, pp 365-386 | |
| 441 | |
| 442 The paper is available at | |
| 443 http://purl.com/STEVEROZEN/papers/rozen-and-skaletsky-2000-primer3.pdf | |
| 444 | |
| 2 | 445 The filtering and assembly of the pal_finder output for Illumina data is performed |
| 446 using a Python utility written by Graeme Fox at the University of Manchester, and which | |
| 447 is included with this tool; this utility uses the BioPython and PANDAseq packages. | |
| 0 | 448 |
| 449 Please kindly acknowledge both this Galaxy tool, the pal_finder and primer3 packages, and | |
| 2 | 450 the utility script and its dependencies if you use it in your work. |
| 0 | 451 </help> |
| 452 <citations> | |
| 453 <!-- | |
| 454 See https://wiki.galaxyproject.org/Admin/Tools/ToolConfigSyntax#A.3Ccitations.3E_tag_set | |
| 455 Can be either DOI or Bibtex | |
| 456 Use http://www.bioinformatics.org/texmed/ to convert PubMed to Bibtex | |
| 457 --> | |
| 458 <citation type="doi">10.1371/journal.pone.0030953</citation> | |
| 459 <citation type="bibtex">@Article{pmid10547847, | |
| 2 | 460 Author="Rozen, S. and Skaletsky, H. ", |
| 0 | 461 Title="{{P}rimer3 on the {W}{W}{W} for general users and for biologist programmers}", |
| 462 Journal="Methods Mol. Biol.", | |
| 463 Year="2000", | |
| 464 Volume="132", | |
| 465 Pages="365--386", | |
| 466 URL="{http://purl.com/STEVEROZEN/papers/rozen-and-skaletsky-2000-primer3.pdf}" | |
| 467 }</citation> | |
| 2 | 468 <citation type="doi">10.1093/bioinformatics/btp163</citation> |
| 469 <citation type="doi">10.1186/1471-2105-13-31</citation> | |
| 0 | 470 </citations> |
| 471 </tool> |