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1 function get_read_counts(anno_dir, outfile, varargin)
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2 %
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3 % -- input --
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4 % anno_dir: directory of genes
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5 % outfile: output file
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6 % varargin: list of BAM files (at least two)
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7
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8 % DESeq paths
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9 global DESEQ_PATH DESEQ_SRC_PATH
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10
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11 % interpreter paths
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12 global INTERPRETER MATLAB_BIN_PATH OCTAVE_BIN_PATH
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13
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14 % SAMTools path
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15 global SAMTOOLS_DIR
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16
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17 %%%% paths
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18 addpath(sprintf('%s/tools', DESEQ_PATH));
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19 addpath(sprintf('%s/mex', DESEQ_PATH));
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20 addpath(sprintf('%s', DESEQ_SRC_PATH));
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21
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22 deseq_config;
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23
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24 %%% read list of replicate groups from variable length argument list
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25 rg_list = cell(1,size(varargin, 2));
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26 file_list = cell();
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27 file_cond_ids = [];
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28 file_rep_ids = [];
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29 for idx = 1:size(varargin, 2)
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30 rg_list(idx) = varargin(idx);
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31 end
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32 idx = strmatch('', rg_list, 'exact');
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33 rg_list(idx) = [];
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34 for idx = 1:length(rg_list),
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35 items = separate(rg_list{idx}, ':');
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36 for idx2 = 1:length(items)
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37 if isempty(deblank(items{idx2})),
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38 continue;
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39 end;
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40 file_list{end + 1} = items{idx2};
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41 file_cond_ids(end + 1) = idx;
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42 file_rep_ids(end + 1) = idx2;
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43 end;
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44 end;
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45 clear idx idx2;
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46
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47 %%%%% adapt to number of input arguments
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48 file_num = length(file_list);
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49 RESULTS = cell(1, file_num);
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50
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51 %%%% get annotation file
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52 load(sprintf('%s', anno_dir));
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53
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54 %%%%% mask overlapping gene regions -> later not counted
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55 [genes] = mask_dubl(genes,0);
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56
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57 %%%% remove genes with no annotated exons or where no
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58 idx = find(arrayfun(@(x)(~isempty(x.exons)*~isempty(x.start)*~isempty(x.stop)), genes));
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59 fprintf('removed %i of %i genes, which had either no exons annotated or lacked a start or stop position\n', size(genes, 2) - size(idx, 2), size(genes, 2))
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60 genes = genes(idx);
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61 clear idx;
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62
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63 %%%% check if genes have field chr_num
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64 if ~isfield(genes, 'chr_num')
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65 chrms = unique({genes(:).chr});
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66 for i = 1:length(genes)
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67 genes(i).chr_num = strmatch(genes(i).chr, chrms, 'exact');
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68 end;
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69 end;
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70
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71 %%%% iterate over all given bam files
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72 for f_idx = 1:file_num
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73 expr1_bam = fullfile('', file_list{f_idx});
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74 STAT = cell(size(genes, 2),1);
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75 for i=1:size(genes,2)
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76 RESULT = cell(1,7);
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77 gene = genes(i);
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78 RESULT{4} = f_idx;
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79 RESULT{1} = gene.name;
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80 if isempty(gene.exons)
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81 RESULT{2} = inf;
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82 RESULT{3} = inf;
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83 RESULT{5} = [inf,inf];
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84 STAT{i} = RESULT;
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85 continue;
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86 elseif or(isempty(gene.start),isempty(gene.stop))
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87 RESULT{2} = inf;
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88 RESULT{3} = inf;
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89 RESULT{5} = [inf,inf];
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90 STAT{i} = RESULT;
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91 continue;
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92 end
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93 if ~isempty(gene.chr_num),
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94 [mask1, read_intron_list] = get_reads(expr1_bam, gene.chr, gene.start, gene.stop, '0');
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95 clear read_intron_list;
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96 else
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97 mask1 = [];
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98 end;
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99
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100 if isempty(mask1)
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101 reads1 = zeros(0,gene.stop-gene.start+1);
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102 else
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103 reads1 = sparse(mask1(1,:)',mask1(2,:)',ones(size(mask1,2),1),max(mask1(1,:)),gene.stop-gene.start+1);
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104 end
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105 if ~isempty(reads1);
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106 [reads1,FLAG] = remove_reads_from_other_genes(reads1,gene);
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107 end
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108 L = size(reads1);
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109 RESULT{2}=[size(reads1,1)]; % number of all reads falling in that gene
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110 EXON_IDX=zeros(1,gene.stop-gene.start+1);
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111 for t=1:size(gene.transcripts,2)
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112 for e=1:size(gene.exons{t},1)
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113 EXON_IDX((gene.exons{t}(e,1)-gene.start+1):(gene.exons{t}(e,2)-gene.start+1))=1;
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114 end
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115 end
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116 reads1 = reads1(sum(reads1(:,find(EXON_IDX)),2)>0,:);
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117 L1 = sum(EXON_IDX);
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118 RESULT{3}=[size(reads1,1)]; % number of reads overlapping to exons
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119 RESULT{5}=[L, L1]; % size of reads1, number of exonic positions
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120 % old and weighted poisson new ,weighted regions reads and
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121 % unexplained reads
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122 clear reads1;
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123 STAT{i} = RESULT;
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124 end;
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125 RESULTS{f_idx} = STAT;
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126 end;
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127
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128 S=size(genes,2);
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129 READCOUNTS_ALL=zeros(S, file_num);
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130 READCOUNTS_EXON=zeros(S, file_num);
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131 LENGTH_ALL=zeros(S,file_num);
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132 LEN_EXON=zeros(S, file_num);
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133
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134 for j=1:file_num,
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135 for i=1:S
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136 T=RESULTS{j}{i};
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137 if isempty(T)
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138 continue
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139 else
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140 READCOUNTS_ALL(i,j)=T{2};
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141 READCOUNTS_EXON(i,j)=T{3};
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142 LENGTH_ALL(i,j)=T{5}(1);
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143 LEN_EXON(i,j)=T{5}(2);
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144 end
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145 end
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146 end
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147
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148 %%%%% write results for all bam files
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149 fid_conditions = fopen(sprintf('%s_CONDITIONS.tab', outfile), 'w');
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150 fid_counts = fopen(sprintf('%s_COUNTS.tab', outfile) ,'w');
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151 fprintf(fid_counts,'gene');
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152 fprintf(fid_conditions, 'file\tcondition\treplicate\n');
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153 for j = 1:length(file_list)
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154 fname = file_list{j} ;
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155 fname = separate(fname, '/');
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156 fname = fname{end};
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157 fname = strrep(fname, '.bam', '') ;
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158 fprintf(fid_counts,'\t%s', fname);
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159 fprintf(fid_conditions, '%s\t%i\t%i\n', fname, file_cond_ids(j), file_rep_ids(j));
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160 end;
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161 fprintf(fid_counts,'\n') ;
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162
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163 for i = 1:size(genes,2)
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164 fprintf(fid_counts,'%s',genes(i).name);
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165 for j = 1:length(file_list),
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166 fprintf(fid_counts,'\t%i', READCOUNTS_EXON(i,j));
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167 end
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168 fprintf(fid_counts,'\n');
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169 end
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170 fclose(fid_counts);
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171 fclose(fid_conditions);
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172 exit;
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