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1 #!/usr/bin/env python
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2
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3 #This is a program to implement the PRADA pipeline "process subsection", based on the work of Rahul Vegesna and Wandaliz Torres-Garcia
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4 #It merely generates a PBS file, depending on the user input entry point (step)
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5 #Author: Siyuan Zheng, szheng2@mdanderson.org
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6 #Copy right belongs to Roel Verhaak's lab from MD Anderson Cancer Center, Department of Bioinformatics and Computational Biology.
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7 #Last revision: 02/17/2014
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8
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9 import subprocess
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10 import os,os.path
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11 import sys
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12 import time
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13 import ioprada
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14 import re
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15
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16 ########################################################################################
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17 args=sys.argv
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18
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19 help_menu='''\nPipeline for RNAseq Data Analaysis - preprocessing pipeline (PRADA).
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20 \t**Usage**:
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21 \tprada-preprocess-bi -conf xx.txt -inputdir .. -sample XX -tag TCGA-XX -platform illumina -step 1_1 -intermediate no -pbs xxx -outdir ... -submit no
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22 \t**Parameters**:
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23 \t-h print help message
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24 \t-step_info print complete steps curated in the module.
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25 \t-inputdir the dir where the input bam or fastq can be found.
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26 \t-sample input sample name. PRADA searches for sample.bam or sample.end1.fastq/sample.end2.fastq, etc, depending on the
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27 \t initiating step number. See step_info for more information.
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28 \t-conf config file for references and parameters. Default is conf.txt in py-PRADA installation folder.
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29 \t-tag a tag to describe the sample, likely sample ID, such as TCGA-LGG-01; no default.
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30 \t-platform only illumina at present (default).
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31 \t-step values: 1_1/2,2_e1/2_1/2/3/4,3_e1/2_1/2,4_1/2,5,6_1/2,7,8; example 2_e1_1; no default.
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32 \t-outdir output dir. Default is the directory where the input bam is.
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33 \t-pbs name for output pbs file and log file. Default (time-stamp) is used if no input.
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34 \t-intermediate values:yes/no; if intermediate files should be kept. Default is not.
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35 \t-submit if submit the job to HPC, default is no. If yes, ppn is set to 12.
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36 \t-v print version information.
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37 '''
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38
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39 steps_info='''
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40 Command orders (sample XX)
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41 step 1_1 --> XX.sorted.bam [sort input bam by name]
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42 step 1_2 --> XX.end1/2.fastq [extract reads from bam]
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43 step 2_e1_1 --> XX.end1.sai [realign end1 reads to composite reference]
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44 step 2_e1_2 --> XX.end1.sam [generate sam]
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45 step 2_e1_3 --> XX.end1.bam [generate bam]
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46 step 2_e1_4 --> XX.end1.sorted.bam [sort bam]
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47 step 2_e2_1 --> XX.end2.sai [realign end2 reads to composite reference]
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48 step 2_e2_2 --> XX.end2.sam [generate sam]
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49 step 2_e2_3 --> XX.end2.bam [generate bam]
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50 step 2_e2_4 --> XX.end2.sorted.bam [sort bam]
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51 step 3_e1_1 --> XX.end1.remapped.bam [remap end1 to genome]
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52 step 3_e1_2 --> XX.end1.remapped.sorted.bam [sort remapped bam]
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53 step 3_e2_1 --> XX.end2.remapped.bam [remap end2 to genome]
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54 step 3_e2_2 --> XX.end2.remapped.sorted.bam [sort remapped bam]
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55 step 4_1 --> XX.paired.bam [pair end1 and end1]
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56 step 4_2 --> XX.paired.sorted.bam [sort paired bam]
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57 step 5 --> XX.withRG.paired.sorted.bam [add read group]
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58 step 6_1 --> XX.orig.csv [prepair recalibration table]
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59 step 6_2 --> XX.withRG.GATKRecalibrated.bam [recalibration]
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60 step 7 --> XX.withRG.GATKRecalibrated.flagged.bam [flag duplication reads]
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61 step 8 --> folder XX for gene expression, QC metrics etc. [generate QC and expression]
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62 '''
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63
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64 if '-h' in args or '-help' in args or len(args)==1:
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65 print help_menu
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66 sys.exit(0)
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67 if '-step_info' in args:
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68 print steps_info
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69 sys.exit(0)
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70 if '-v' in args:
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71 import version
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72 print version.version
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73 sys.exit(0)
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74
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75 if '-sample' not in args:
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76 sys.exit('ERROR: Sample name is needed')
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77 if '-step' not in args:
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78 sys.exit('ERROR: Step number is needed')
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79 if '-tag' not in args:
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80 sys.exit('ERROR: A tag is needed')
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81
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82 i=args.index('-sample')
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83 sample=args[i+1]
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84 if '-inputdir' not in args:
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85 inputpath=os.path.abspath('./')
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86 else:
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87 i=args.index('-inputdir')
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88 inputpath=args[i+1]
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89 bampath=os.path.abspath(inputpath)+'/%s.bam'%sample
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90 fq1path=os.path.abspath(inputpath)+'/%s.end1.fastq'%sample
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91 fq2path=os.path.abspath(inputpath)+'/%s.end2.fastq'%sample
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92
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93 if '-outdir' not in args:
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94 outpath=os.path.dirname(bampath)
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95 else:
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96 i=args.index('-outdir')
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97 outpath=os.path.abspath(args[i+1])
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98 if not os.path.exists(outpath):
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99 os.mkdir(outpath)
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100
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101 #bam=os.path.basename(bampath)
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102 #sample=bam[:-4]
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103 i=args.index('-step')
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104 step=args[i+1]
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105 i=args.index('-tag')
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106 tag=args[i+1]
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107
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108 prada_path=os.path.dirname(os.path.abspath(__file__)) ####
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109 ref_search_path=[prada_path,os.getcwd()] #search path for ref file if not specified in command line
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110
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111 if '-conf' in args:
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112 i=args.index('-conf')
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113 reffile=args[i+1]
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114 if os.path.exists(reffile):
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115 pass
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116 else:
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117 for pth in ref_search_path:
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118 new_reffile='%s/%s'%(pth, os.path.basename(reffile))
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119 if os.path.exists(new_reffile):
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120 reffile=new_reffile
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121 break
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122 else:
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123 sys.exit('ERROR: conf file %s not found'%reffile)
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124 else:
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125 reffile='%s/conf.txt'%prada_path
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126 if not os.path.exists(reffile):
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127 sys.exit('ERROR: No default conf.txt found and none specified')
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128
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129 if '-platform' in args:
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130 i=args.index('-platform')
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131 plat=args[i+1]
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132 else:
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133 plat='illumina'
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134 if '-submit' in args:
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135 i=args.index('-submit')
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136 submit=args[i+1]
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137 else:
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138 submit='no'
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139 if '-intermediate' in args:
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140 i=args.index('-intermediate')
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141 keepmed=args[i+1]
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142 else:
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143 keepmed='False'
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144
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145 if keepmed in ['False','FALSE','false','F','NO','No','no','n']:
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146 keepmed_flag='no'
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147 elif keepmed in ['True','TRUE','true','T','YES','Yes','yes','y']:
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148 keepmed_flag='yes'
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149 else:
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150 sys.exit('ERROR: -intermediate value not recognized')
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151
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152 if '-pbs' in args:
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153 i=args.index('-pbs')
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154 docstr=args[i+1]
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155 else:
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156 a=time.ctime().split()
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157 b=time.time()
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158 timestamp='_'.join([a[-1],a[1],a[2]])+'.'+str(b)
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159 docstr='prada_prep_'+timestamp
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160 logfilename=docstr+'.log'
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161 pbsfilename=docstr+'.pbs'
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162 pbspath=outpath+'/'+pbsfilename
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163 logpath=outpath+'/'+logfilename
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164 ########################################################################################
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165
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166 ########################################################################################
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167 #underlying utilities, automatically detected
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168 samtools='%s/tools/samtools-0.1.16/samtools'%prada_path
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169 bwa='%s/tools/bwa-0.5.7-mh/bwa'%prada_path
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170 gatk='%s/tools/GATK/'%prada_path
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171 picard='%s/tools/Picard/'%prada_path
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172 seqc='%s/tools/RNA-SeQC_v1.1.7.jar'%prada_path
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173 #Default uses 12 nodes in HPC
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174 #########################################################################################
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175
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176 #########################################################################################
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177 #reference files
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178 refdict=ioprada.read_conf(reffile)
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179 genome_gtf=refdict['--REF--']['genome_gtf']
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180 compdb_fasta=refdict['--REF--']['compdb_fasta']
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181 compdb_map=refdict['--REF--']['compdb_map']
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182 genome_fasta=refdict['--REF--']['genome_fasta']
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183 dbsnp_vcf=refdict['--REF--']['dbsnp_vcf']
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184 select_tx=refdict['--REF--']['select_tx']
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185 pat=re.compile('ppn=(\d*)')
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186 parallel_n=pat.search(refdict['--PBS--']['-l']).groups()[0]
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187 #########################################################################################
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188
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189 #########################################################################################
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190 #pipeline command lines.
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191 ##Cleaning up steps: if -intermediate is yes, none will be executed.
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192 step_1_1_cmd=['%s sort -n -m 1000000000 %s %s.sorted'%(samtools,bampath,sample)]
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193 post_1_1_clean=[]
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194 step_1_2_cmd=['java -Xmx8g -jar %s/SamToFastq.jar INPUT=%s.sorted.bam FASTQ=%s.end1.fastq SECOND_END_FASTQ=%s.end2.fastq INCLUDE_NON_PF_READS=true VALIDATION_STRINGENCY=SILENT TMP_DIR=tmp/'%(picard,sample,sample,sample)]
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195 post_1_2_clean=['rm -f %s.sorted.bam'%sample]
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196 alnparams=' '.join([' '.join(x) for x in refdict['--BWA aln--'].items()])
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197 samseparams=' '.join([' '.join(x) for x in refdict['--BWA samse--'].items()])
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198 if step == '2_e1_1' or step == '2_e2_1':
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199 step_2_e1_1_cmd=['%s aln %s %s %s > %s.end1.sai'%(bwa,alnparams,compdb_fasta,fq1path,sample)]
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200 post_2_e1_1_clean=[]
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201 step_2_e1_2_cmd=['%s samse -s %s %s %s.end1.sai %s > %s.end1.sam'%(bwa,samseparams,compdb_fasta,sample,fq1path,sample)]
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202 post_2_e1_2_clean=['rm -f %s.end1.sai'%sample]
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203 step_2_e2_1_cmd=['%s aln %s %s %s > %s.end2.sai'%(bwa,alnparams,compdb_fasta,fq2path,sample)]
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204 post_2_e2_1_clean=[]
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205 step_2_e2_2_cmd=['%s samse -s %s %s %s.end2.sai %s > %s.end2.sam'%(bwa,samseparams,compdb_fasta,sample,fq2path,sample)]
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206 post_2_e2_2_clean=['rm -f %s.end2.sai'%sample]
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207 else:
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208 step_2_e1_1_cmd=['%s aln %s %s %s.end1.fastq > %s.end1.sai'%(bwa,alnparams,compdb_fasta,sample,sample)]
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209 post_2_e1_1_clean=[]
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210 step_2_e1_2_cmd=['%s samse -s %s %s %s.end1.sai %s.end1.fastq > %s.end1.sam'%(bwa,samseparams,compdb_fasta,sample,sample,sample)]
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211 post_2_e1_2_clean=['rm -f %s.end1.sai'%sample,'rm -f %s.end1.fastq'%sample]
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212 step_2_e2_1_cmd=['%s aln %s %s %s.end2.fastq > %s.end2.sai'%(bwa,alnparams,compdb_fasta,sample,sample)]
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213 post_2_e2_1_clean=[]
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214 step_2_e2_2_cmd=['%s samse -s %s %s %s.end2.sai %s.end2.fastq > %s.end2.sam'%(bwa,samseparams,compdb_fasta,sample,sample,sample)]
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215 post_2_e2_2_clean=['rm -f %s.end2.sai'%sample,'rm -f %s.end2.fastq'%sample]
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216 step_2_e1_3_cmd=['%s view -bS -o %s.end1.bam %s.end1.sam'%(samtools,sample,sample)]
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217 post_2_e1_3_clean=['rm -f %s.end1.sam'%sample]
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218 step_2_e1_4_cmd=['%s sort -n -m 1000000000 %s.end1.bam %s.end1.sorted'%(samtools,sample,sample)]
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219 post_2_e1_4_clean=['rm -f %s.end1.bam'%sample]
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220 step_2_e2_3_cmd=['%s view -bS -o %s.end2.bam %s.end2.sam'%(samtools,sample,sample)]
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221 post_2_e2_3_clean=['rm -f %s.end2.sam'%sample]
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222 step_2_e2_4_cmd=['%s sort -n -m 1000000000 %s.end2.bam %s.end2.sorted'%(samtools,sample,sample)]
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223 post_2_e2_4_clean=['rm -f %s.end2.bam'%sample]
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224 step_3_e1_1_cmd=['java -Djava.io.tmpdir=tmp/ -cp %s/RemapAlignments.jar -Xmx8g org.broadinstitute.cga.tools.gatk.rna.RemapAlignments M=%s IN=%s.end1.sorted.bam OUT=%s.end1.remapped.bam R=%s REDUCE=TRUE'%(gatk,compdb_map,sample,sample,genome_fasta)]
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225 post_3_e1_1_clean=['rm -f %s.end1.sorted.bam'%sample]
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226 step_3_e1_2_cmd=['%s sort -n -m 1000000000 %s.end1.remapped.bam %s.end1.remapped.sorted'%(samtools,sample,sample)]
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227 post_3_e1_2_clean=['rm -f %s.end1.remapped.bam'%sample]
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228 step_3_e2_1_cmd=['java -Djava.io.tmpdir=tmp/ -cp %s/RemapAlignments.jar -Xmx8g org.broadinstitute.cga.tools.gatk.rna.RemapAlignments M=%s IN=%s.end2.sorted.bam OUT=%s.end2.remapped.bam R=%s REDUCE=TRUE'%(gatk,compdb_map,sample,sample,genome_fasta)]
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229 post_3_e2_1_clean=['rm -f %s.end2.sorted.bam'%sample]
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230 step_3_e2_2_cmd=['%s sort -n -m 1000000000 %s.end2.remapped.bam %s.end2.remapped.sorted'%(samtools,sample,sample)]
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231 post_3_e2_2_clean=['rm -f %s.end2.remapped.bam'%sample]
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232 step_4_1_cmd=['java -Djava.io.tmpdir=tmp/ -Xmx8g -jar %s/PairMaker.jar IN1=%s.end1.remapped.sorted.bam IN2=%s.end2.remapped.sorted.bam OUTPUT=%s.paired.bam TMP_DIR=tmp/'%(gatk,sample,sample,sample)]
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233 post_4_1_clean=['rm -f %s.end1.remapped.sorted.bam'%sample,'rm -f %s.end2.remapped.sorted.bam'%sample]
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234 step_4_2_cmd=['%s sort -m 1000000000 %s.paired.bam %s.paired.sorted'%(samtools,sample,sample)]
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235 post_4_2_clean=['rm -f %s.paired.bam'%sample]
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236 step_5_cmd=['java -Xmx8g -jar %s/AddOrReplaceReadGroups.jar I=%s.paired.sorted.bam O=%s.withRG.paired.sorted.bam RGLB=%s RGPL=%s RGPU=%s RGSM=%s'%(picard,sample,sample,tag,plat,tag,tag)]
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237 post_5_clean=['rm -f %s.paired.sorted.bam'%sample]
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238 step_6_1_cmd=['%s index %s.withRG.paired.sorted.bam'%(samtools,sample),'java -Xmx8g -jar %s/GenomeAnalysisTK.jar -l INFO -R %s --default_platform %s --knownSites %s -I %s.withRG.paired.sorted.bam --downsample_to_coverage 10000 -T CountCovariates -cov ReadGroupCovariate -cov QualityScoreCovariate -cov CycleCovariate -cov DinucCovariate -nt %s -recalFile %s.orig.csv'%(gatk,genome_fasta,plat,dbsnp_vcf,sample,parallel_n,sample)]
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239 post_6_1_clean=[]
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240 step_6_2_cmd=['java -Xmx8g -jar %s/GenomeAnalysisTK.jar -l INFO -R %s --default_platform %s -I %s.withRG.paired.sorted.bam -T TableRecalibration --out %s.withRG.GATKRecalibrated.bam -recalFile %s.orig.csv'%(gatk,genome_fasta,plat,sample,sample,sample)]
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241 post_6_2_clean=['rm -f %s.withRG.paired.sorted.bam'%sample,'rm -f %s.withRG.paired.sorted.bam.bai'%sample,'rm -f %s.orig.csv'%sample]
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242 step_7_cmd=['java -Xmx8g -jar %s/MarkDuplicates.jar I=%s.withRG.GATKRecalibrated.bam O=%s.withRG.GATKRecalibrated.flagged.bam METRICS_FILE=%s.Duplicates_metrics.txt VALIDATION_STRINGENCY=SILENT TMP_DIR=tmp/'%(picard,sample,sample,sample),'%s index %s.withRG.GATKRecalibrated.flagged.bam'%(samtools,sample)]
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243 post_7_clean=['rm -f %s.withRG.GATKRecalibrated.bam'%sample,'rm -f %s.withRG.GATKRecalibrated.bai'%sample,'rm -f %s.Duplicates_metrics.txt'%sample]
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244 step_8_cmd=["java -Xmx8g -jar %s -ttype 2 -t %s -r %s -s '%s|%s.withRG.GATKRecalibrated.flagged.bam|Disc' -o %s/"%(seqc,genome_gtf,genome_fasta,sample,sample,sample)]
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245 post_8_clean=[]
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246
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247 cmdset=[]
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248 for item in globals().keys():
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249 if item.endswith('_cmd'):
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250 cmdset.append(item)
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251 cmdset.sort()
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252
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253 #########################################################################################
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254 #write PBS file
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255
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256 ###############Entry Point
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257 try:
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258 cmd_entry=cmdset.index('step_'+step+'_cmd') ##entry point
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259 except ValueError:
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260 sys.exit('ERROR: STEP not recognized')
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261
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262 def _parsecmd(cmd):
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263 '''pase cmd str for step information'''
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264 info=cmd.split('_')
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265 cstep='_'.join(info[1:-1])
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266 return cstep
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267
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268 ###########################
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269 ##headers
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270 outfile=open(pbspath,'w')
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271 outfile.write('#! /bin/sh\n')
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272 outfile.write('#PBS -V\n')
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273 outfile.write('#PBS -N %s\n'%tag)
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274 outfile.write('#PBS -j oe\n')
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275 outfile.write('#PBS -o %s\n'%logpath)
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276 for item in refdict['--PBS--']:
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277 outfile.write('#PBS %s %s\n'%(item,refdict['--PBS--'][item]))
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278 outfile.write('#PBS -d %s\n'%outpath)
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279
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280 #commands
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281 outfile.write('echo "Job start: `date`"\n')
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282
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283 for i in range(cmd_entry,len(cmdset)):
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284 cmd=cmdset[i]
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285 cstep=_parsecmd(cmd)
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286 clean_flag=1
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287 outfile.write('echo "step %s start: `date`"\n'%cstep)
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288 outfile.write('if\n')
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289 outfile.write('\t%s\n'%('\n'.join(eval(cmd))))
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290 outfile.write('then\n')
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291 outfile.write('\techo "step %s done: `date`"\n'%cstep)
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292 outfile.write('else\n')
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293 outfile.write('\techo "step %s ERROR"\n'%cstep)
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294 outfile.write('\texit\n')
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295 outfile.write('fi\n')
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296
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297 if keepmed_flag=='yes': #if so, keep files by force
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298 clean_flag=0
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299
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300 if clean_flag==1:
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301 outfile.write('\n'.join(eval('post_%s_clean'%cstep)))
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302 outfile.write('\n')
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303
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304 outfile.write('echo "PIPELINE FINISHED"\n')
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305 outfile.close()
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306 ######################################################################
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307
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308 if submit in ['False','FALSE','false','F','NO','No','no','n']:
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309 jid='Not_Submitted'
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310 logpath='None'
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311 elif submit in ['True','TRUE','true','T','YES','Yes','yes','y']:
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312 cmdstr='qsub %s'%pbspath
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313 cmd=cmdstr.split()
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314 cmdout=subprocess.Popen(cmd,stdout=subprocess.PIPE,stderr=subprocess.STDOUT)
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315 jid=cmdout.stdout.read().strip() ##JOB ID
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316 else:
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317 sys.exit('ERROR: submit parameter not recognized')
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318
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319 print '#!#%s'%tag
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320 print 'BAM\t%s'%bampath
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321 print 'Entry\t%s'%step
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322 print 'PBS\t%s'%pbspath
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323 print 'JOB\t%s'%jid
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324 print 'LOG\t%s'%logpath
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325
|
