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date | Tue, 07 Nov 2017 12:28:31 -0500 |
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#!/usr/bin/env perl #============================================================================================== # Script to output statistsics and histograms for reads and contigs/isotigs # Outputs include: # Mean, N50, StdDev or reads or contig lengths, # Mean and Modal read or contig lengths. # Number of reads or contigs > 1 kb in length # Summed contig length (by number of contigs, in sorted order) # Histogram of read or contig lengths, # Graph of sums of read-lengths # File of reads or contigs sorted by read or contig length # Test for mono/di-nucelotide repeats # Randomly selected reads or contigs # Needs gnuplot installed to create the histograms: # On Fedora/Redhat linux: sudo yum install gnuplot # On Ubuntu/Debian: sudo apt-get install gnuplot # Uses a linux pipe to call gnu-plot directly, rather than as a separate shell script. # Original written by Sujai Kumar, 2008-09-05 University of Edinburgh # Modified by Stephen: 29-Apr-2009: # Last changed by Stephen: 9-Aug-2010 # Usage: fasta_summary.pl -i infile.fasta -o process_reads -t read OR contig OR isotig (to use 'read' or 'contig' or 'isotig' in the output table & graphs. Isotig is for 'runAssembly -cdna ...' output file '454Isotigs.fna') [-r 1 to indicate count simple nucleotide repeats] [-n number of random reads to output] [-c cutoff_length] [-l 1 to indicate output the longest read] [-f (s or t or w) for spacer, tab or wiki format table output.] # Note: The parameters above in the [] are optional. # eg: fasta_summary.pl -i myfile.fasta -o process_reads -t read # Where: # -i reads or contigs as input, in fasta format. # -o output_dir (created if it doesn't exist) # -t read, contig or isotig # Gives back # - N50 # - num of contigs > 1 kb # - num of contigs # - Read or Contig Histogram and graphs. # - Summed contig length (by number of contigs, in sorted order) #============================================================================================== use strict; use warnings; use Getopt::Long; my $infile; my $output_dir; my $type='read'; # Defaults to 'read' at present my $repeats=1; my $num_random_reads_to_output=0; my $cutoff_length=-1; # -1 means won't check this cutoff my $longest_read=-1; # -1 mean's don't output the sequence for the longest read. my $doCommify=1; # Outputs statistics numbers in format: 9,999,999 my $format="t"; # "s"=spaces between columns, "t"=tabs between columns, "w"=wiki '||' and '|'. my $bucket1=0; # For optional exact length histogram distribution as asked for by JH. if ($#ARGV==-1) {die " Usage: fasta_summary.pl -i infile.fasta -o output_dir -t ( read | contig | isotig ) [ -r 0 ] [ -n num_reads ] [ -c cutoff_length ] [ -l 1 ] [ -d 0 ] [ -f (w | t ) ] [ -bucket1 ] where: -i or -infile infile.fasta : input fatsa file of raeds, contigs or isotigs, -o or -output_dir output_directory : directory to put output stats and graphs into. -t or -type (read or contig or isotig) : for displaying the graph title, where type is 'read' or 'contig' or 'isotig'. -r or -repeats 0 or 1 : 1=count number of reads that contain over 70% simple mono-nucleotide and di-nucleotide repeat bases; 0=don't count. -n or -number num_reads : For outputting specified number of randomly selected reads or contigs. -c or -cutoff cutoff_length : Give a number of reads to do extra analysis (calculating again the number of reads and number of bases in reads above this length) -l or -longest 0 or 1 : 1=Output the longest read; 0= don't output the longest read -d or -doCommify 0 or 1 : Output numbers formatted with commas to make easier to read: 0=no commas, default=1 -f or -format w or t : w=wiki_format (ie. table with || and | for column dividers), t=tabs between column symbols for the wiki pages, default is spaces between columns. -b or -bucket1 : To also output histogram file of exact read lengths (ie. bucket size of 1) eg: For 454 reads: fasta_summary.pl -i RawReads.fna -o read_stats -t read For 454 isotigs: fasta_summary.pl -i 454Isotigs.fna -o isotig_stats -t isotig ";} GetOptions ( "infile=s" => \$infile, "output_dir=s" => \$output_dir, "type=s" => \$type, ## type is 'read' or 'contig' or 'isotig' - for displaying the graph title "repeats=i" => \$repeats, # To count simple repeats "number=i" => \$num_random_reads_to_output, # For outputting specified number of random reads "cutoff=i" => \$cutoff_length, # Give a number of reads to do extra analysis (calculating again the number of reads and number of bases in reads above this length) "longest=i" => \$longest_read, # Output the longest read. "doCommify=i" => \$doCommify, # Output numbers formatted with commas to make easier to read: 0=no commas, default=1 "format=s" => \$format, # "w"=wiki_format (ie. table with || and | for column dividers), "t"=tabs between column symbols for the wiki pages, default is spaces between columns. "bucket1" => \$bucket1, # To also output histogram file of exact read lengths (ie. bucket size of 1) ); if ($#ARGV>-1) {die "Unused options specified: @ARGV\n";} if ( (! defined $infile) || ($infile eq '') ) {die "\nPlease give input fasta file, preceeded by the '-i' option\n\n";} if ( (! defined $output_dir) || ($output_dir eq '') ) {die "Please give output_directory, preceeded by the '-o' option\n\n";} if ( (! defined $type) || (($type ne 'contig') && ($type ne 'read') && ($type ne 'isotig')) ) {die "ERROR: On commandline: -t type must be 'contig' or 'read' or 'isotig'\n\n";} my ($L,$M,$R, $Lh,$Mh,$Rh, $Lhnewline,$Mhnotab); if ($format eq 's') {($L,$M,$R, $Lh,$Mh,$Rh, $Lhnewline,$Mhnotab)=('',' ','', '',' ','', "\n",'');} elsif ($format eq 't') {($L,$M,$R, $Lh,$Mh,$Rh, $Lhnewline,$Mhnotab)=("\t","\t",'', "","\t",'', "\n",'');} # There is correctly a tab for the $L, but not the $Lh. elsif ($format eq 'w') {($L,$M,$R, $Lh,$Mh,$Rh, $Lhnewline,$Mhnotab)=('| ',' | ',' |', '|| ',' || ',' ||', '|| ',' || ');} else {die "\nInvalid output format code: '$format'. Should be 's', 't' or 'w'.\n\n";} ### create output_dir if it doesn't exist if (-d $output_dir) { print STDERR " Directory '$output_dir' exists, so the existing fasta_summary.pl output files will be overwritten\n"; } else { mkdir $output_dir; print STDERR " Directory '$output_dir' created\n"; } my $gc_count = 0; #--------------- Read in contigs from fasta file ------------------- open INFILE, "<$infile" or die "Failed to open file: '$infile' : $!"; open STATS, ">$output_dir/stats.txt" or die "Failed to open $output_dir/stats.txt: '' : $!"; my $header = <INFILE>; if (! defined($header) ) {print "\n** ERROR: First line of input fasta file is undefined - so file must be empty **\n\n"; print STATS "No sequences found\n"; exit 1;} if ($header!~/^>/) {print "\nERROR: First line of input fasta file should start with '>' BUT first line is: '$header'\n"; print STATS "No sequences found\n"; exit 1;} my $seq = ""; my @sequences; while (<INFILE>) { if (/^>/) { push @sequences, [length($seq), $header, $seq]; $header = $_; $seq = ""; } else { chomp; $gc_count += tr/gcGC/gcGC/; $seq .= $_; } } push @sequences, [length($seq), $header, $seq]; close INFILE; if ($#sequences==-1) {print "\nERROR: There are zero sequences in the input file: $infile\n\n"; print STATS "No sequences found\n"; exit 1;} my $all_contigs_length=0; foreach (@sequences) {$all_contigs_length += $_->[0];} if ($all_contigs_length==0) {print "\nERROR: Length of all contigs is zero\n\n"; exit 2;} # Find number and number of bases in reads greater than the optional cut-off length given at command-line. my $num_reads_above_cutoff=0; my $num_of_bases_in_reads_above_cutoff=0; if ($cutoff_length>0) { foreach (@sequences) { if ($_->[0]>=$cutoff_length) {$num_of_bases_in_reads_above_cutoff+= $_->[0]; $num_reads_above_cutoff++;} } } #--------------- Gather Plots Data, Find N50, Print sorted contig file ------------------- my $summed_contig_length = 0; my @summed_contig_data; # <-- For graph of summed length (in number of bases) versus number of contigs. my @summed_contig_data_contigLens; # <-- Added by SJBridgett to get graph of summed contig length versus min. contig length included (ie. X-axis is sort of inverse of above) my $contig1k_count = 0; my $contig1k_length = 0; open SORTED, ">$output_dir/sorted_contigs.fa" or die $!; # top row in stats file #print STATS "N50\nMax contig size\nNumber of bases in contigs\nNumber of contigs\nNumber of contigs >=1kb\nNumber of contigs in N50\nNumber of bases in contigs >=1kb\nGC Content of contigs\n"; my $N50size=-1; my $N50_contigs = 0; my @sorted_by_contig_length = sort {$b->[0] <=> $a->[0]} @sequences; ### variables and initialization for histogram (stored in @bins) my $max = $sorted_by_contig_length[0][0]; my $mean= $all_contigs_length/($#sequences+1); # <-- Added by Stephen Bridgett. Note: as $# gives the highest index number, so add 1 as arrays are zero-based. # Calculate standard deviation my $sumsquares = 0; foreach (@sequences) {$sumsquares += ($_->[0] - $mean) ** 2;} # <-- Taken from John's "mean_fasta_length.pl" script. my $stddev = ( $sumsquares/($#sequences+1) ) ** 0.5; my $min = 0; # Aim for approximately 100 bins, so my $bin_size=1; my $min_max_range=$max - $min; # $bin_size = ($min_max_range)/(99); # (99 is 100-1) so 1000/100 if ($min_max_range>=100000000) {$bin_size=1000000;} elsif ($min_max_range>=10000000) {$bin_size=100000;} elsif ($min_max_range>=1000000) {$bin_size=10000;} elsif ($min_max_range>=100000) {$bin_size=1000;} elsif ($min_max_range>=10000) {$bin_size=100;} else {$bin_size=10;} # elsif ($min_max_range>=1000) {$bin_size=10;} #elsif ($min_max_range>=100) {$bin_size=1;} #elsif ($min_max_range>=10) {} #elsif ($min_max_range>=1) {} # WAS: my $bin_size = ($type eq 'contig') ? 1000 : 10; my @bins; $#bins = int(($min_max_range)/$bin_size) + 1; # <-- Set the bins array size. foreach (@bins) {$_ = 0}; foreach (@sorted_by_contig_length) { my $curr_contig_length = $_->[0]; push @summed_contig_data_contigLens, $curr_contig_length; # <-- added by Stephen. $bins[int(($curr_contig_length + 1 - $min)/$bin_size)]++; $summed_contig_length += $curr_contig_length; push @summed_contig_data, $summed_contig_length; ### sorted contigs file print SORTED $_->[1] . $_->[2] . "\n"; if ($curr_contig_length >= 1000) { $contig1k_count++; $contig1k_length += $curr_contig_length; } $N50_contigs++ unless ($N50size>-1); # Was unless $N50_found if ($summed_contig_length > ($all_contigs_length / 2) and $N50size == -1) { $N50size = $curr_contig_length; } } if ($bucket1!=0) { =pod # This firsdt method works and agress with the second, but the lengths are in reverse order, at the @sorted_by_contig_length array was sorted with longest contig first. open BUCKET1, ">$output_dir/lengths_hist1.txt" or die "Failed to open file '$output_dir/lengths_hist1.txt' : $!\n"; print BUCKET1 "Length\tFrequency\n"; my $len=-1; my $count=0; foreach (@sorted_by_contig_length) { if ( $len != $_->[0] ) {if ($len>-1) {print BUCKET1 "$len\t$count\n";} $len=$_->[0]; $count=0;} $count++; } if ($len>-1) {print BUCKET1 "$len\t$count\n";} # Print length of final length grouping. close BUCKET1; =cut open BUCKET1, ">$output_dir/lengths_hist1_with_zeros.txt" or die "Failed to open file '$output_dir/lengths_hist1_with_zeros.txt' : $!\n"; print BUCKET1 "Length\tFrequency\n"; my @bucket=(); # To check the result by using array. foreach (@sequences) { my $len=$_->[0]; if (defined $bucket[$len]) {$bucket[$len]++;} else {$bucket[$len]=1;} } for (my $i=0; $i<=$#bucket; $i++) # for (my $i=$#bucket; $i>=0; $i--) # <-- for reverse order { if (defined $bucket[$i]) {print BUCKET1 "$i\t$bucket[$i]\n";} else {print BUCKET1 "$i\t0\n";} # Can uncomment this later if don't want zeros in the output. } close BUCKET1; } my $type_plural=$type.'s'; print STATS $Lh."Statistics for $type lengths:".$Mhnotab.$Rh."\n"; print STATS $L."Min $type length:".$M.&commify_if($sorted_by_contig_length[$#sequences][0],$doCommify).$R."\n"; print STATS $L."Max $type length:".$M.&commify_if($max,$doCommify).$R."\n"; printf STATS $L."Mean %s length:".$M."%.2f".$R."\n", $type,$mean; # <-- Added by Stephen Bridgett, April 2009. printf STATS $L."Standard deviation of %s length:".$M."%.2f".$R."\n", $type,$stddev; ## <-- Added by Stephen Bridgett, May 2009. print STATS $L."Median $type length:".$M.&commify_if($sorted_by_contig_length[int($#sequences/2)][0],$doCommify).$R."\n"; print STATS $L."N50 $type length:".$M.&commify_if($N50size,$doCommify).$R."\n"; print STATS $Lhnewline."Statistics for numbers of $type_plural:".$Mhnotab.$Rh."\n"; print STATS $L."Number of $type_plural:".$M.&commify_if($#sequences+1,$doCommify).$R."\n"; print STATS $L."Number of $type_plural >=1kb:".$M.&commify_if($contig1k_count,$doCommify).$R."\n"; print STATS $L."Number of $type_plural in N50:".$M.&commify_if($N50_contigs,$doCommify).$R."\n"; print STATS $Lhnewline."Statistics for bases in the $type_plural:".$Mhnotab.$Rh."\n"; print STATS $L."Number of bases in all $type_plural:".$M.&commify_if($all_contigs_length,$doCommify).$R."\n"; print STATS $L."Number of bases in $type_plural >=1kb:".$M.&commify_if($contig1k_length,$doCommify).$R."\n"; printf STATS $L."GC Content of %s:".$M."%.2f %%".$R."\n", $type_plural,(100*$gc_count/$all_contigs_length); if ($cutoff_length>0) { print STATS $Lhnewline."Statistics for $type_plural >= $cutoff_length bp in length:".$Mhnotab.$Rh."\n"; print STATS $L."Number of $type_plural >= $cutoff_length bp:".$M.&commify($num_reads_above_cutoff,$doCommify).$R."\n"; print STATS $L."\tNumber of bases in $type_plural >= $cutoff_length bp:".$M.&commify($num_of_bases_in_reads_above_cutoff,$doCommify).$R."\n"; } if ($repeats==1) {&countRepeats();} print STATS "\n"; # Output random selection of reads if requested on commandline: my $fastaLineLen=60; # <-- The line length used for 454 sffinfo output, but could use a value read from input file (but be careful not to read a short line) if ($num_random_reads_to_output>0) { my @randlist; if ($num_random_reads_to_output<($#sequences+1)) { print STATS "\nSome randomly selected reads:\n\n"; @randlist= &getListOfRandomNumbers($#sequences, $num_random_reads_to_output); # Don't use ($#sequences + 1), just ($#sequences) otherwise would be outside the array. } else # Just print all the sequences: { print STATS "\nAll ".($#sequences+1)." reads:\n\n"; for (my $i=0;$i<=$#sequences;$i++) {push @randlist,$i;} } &print_sequences(\@randlist) } # Print the longest read: if ($longest_read>0) { my $length_of_longest_read=-1; my @longest_read=(); my $i=0; foreach (@sequences) { if ($_->[0]>$length_of_longest_read) {$length_of_longest_read=$_->[0]; $longest_read[0]=$i;} $i++; } if ($length_of_longest_read>0) {print STATS "\nLongest read:\n"} &print_sequences(\@longest_read); } =pod print STATS "\n$type\tSummed\nlength\tlength\n"; # <-- Added by Stephen Bridgett, but better to produce a graph instead. my $i=0; foreach (@summed_contig_data) { # print STATS $sorted_by_contig_length[$i]->[0]."\t".$summed_contig_data_contigLens[$i]."\t".$_."\t".$summed_contig_data[$i]."\n"; print STATS $sorted_by_contig_length[$i]->[0]."\t".$_."\n"; $i++; } =cut open SUMMED, ">$output_dir/summed_contig_lengths.dat" or die $!; print SUMMED join "\n",@summed_contig_data; close SUMMED; open HISTOGRAMBINS, ">$output_dir/histogram_bins.dat" or die $!; my $bin_size_counter = 0; foreach (@bins) { print HISTOGRAMBINS eval($bin_size_counter++ * $bin_size + $bin_size/2) . "\t$_\n"; } close HISTOGRAMBINS; # Graph of cumulative (summed) number of reads on y-axis, versus length of read (decending order) on x-axis open SUMREAD_READLEN, ">$output_dir/sum_reads_vs_read_len.dat" or die $!; #my $read_counter= 0; my $read_counter= $#sorted_by_contig_length+1; foreach (@sorted_by_contig_length) { # $read_counter++; $read_counter--; print SUMREAD_READLEN "$_->[0]\n"; # $read_counter } close SUMREAD_READLEN; my $properType=ucfirst($type); # Makes the first letter an upper case letter, ie. 'Config' or 'Read' #if ($type eq 'contig') # { # print the outcome of the gnu_plot as may have a write permissions error sometimes. my $YHistogramScaleType = ($type eq 'read') ? '' : 'log y'; # Not using log scale for reads, just for contig/isotigs. &plot_graph('histogram', "$output_dir/histogram_bins.dat", "Histogram of $type lengths", "$properType length", "Number of $type_plural", '0.9', $YHistogramScaleType); &plot_graph('line', "$output_dir/summed_contig_lengths.dat", "Summed $type lengths", "Number of $type_plural", "Summed $type length in bases", '0.9', ''); &plot_graph('line', "$output_dir/sum_reads_vs_read_len.dat", "X-axis gives the Number of $type_plural that are greater than the $properType-length given on the Y-axis", "$properType length", "Cummulative number of $type_plural", '0.9', ''); =pod # print `gnuplot_histogram.sh $output_dir/histogram_bins.dat`; &plot_graph("$output_dir/summed_contig_lengths.dat", 'Summed contig lengths', 'Number of contigs', 'Summed contig length in bases', '0.9', ''); # print `gnuplot_summedcontigs.sh $output_dir/summed_contig_lengths.dat`; &plot_graph('line', "$output_dir/sum_reads_vs_read_len.dat", 'X-axis gives the Number of contigs that are greater than the Contig-length given on the Y-axis', 'Contig length', 'Cummulative number of contigs', '0.9', ''); # print `gnuplot_sum_contig_vs_contig_len.sh $output_dir/sum_reads_vs_read_len.dat`; } elsif ($type eq 'read') { # print `gnuplot_readshistogram_logY.sh $output_dir/histogram_bins.dat`; # There's also optionally a "...._linearY.sh" &plot_graph('line', "$output_dir/summed_contig_lengths.dat",'Summed read lengths', 'Number of reads', 'Summed read length in bases', '0.9', ''); # print `gnuplot_summedreads.sh $output_dir/summed_contig_lengths.dat`; &plot_graph('line', "$output_dir/sum_reads_vs_read_len.dat", 'X-axis gives the Number of reads that are greater than the Read-length given on the Y-axis', 'Read length', 'Cummulative number of reads', '0.9', ''); # print `gnuplot_sum_reads_vs_read_len.sh $output_dir/sum_reads_vs_read_len.dat`; } else {die "\n** ERROR: Invalid type='$type' **\n\n";} =cut close SORTED; close STATS; # Use pipe to plot directly with gnuplot, rather than calling a separate shell script: # http://www.vioan.ro/wp/2008/09/30/calling-gnuplot-from-perl/ # http://forums.devshed.com/perl-programming-6/plotting-with-gnuplot-within-perl-script-549682.html # Another option is the perl module: GnuplotIF: http://lug.fh-swf.de/perl/GnuplotIF.html OR: http://lug.fh-swf.de/perl/ # PlPlot: Perl: http://search.cpan.org/~dhunt/PDL-Graphics-PLplot-0.47/plplot.pd # http://plplot.sourceforge.net/ # dislin: http://www.mps.mpg.de/dislin/overview.html # MathGL: http://mathgl.sourceforge.net/index.html sub plot_graph { # Plots a histogram or xy-line graph # Parameters: GraphType (histogram/line) DataFile, Title, X-label, Y-label, Y-range # Graphfile should end with '.png' # The $yloglinear is 'log y' for log, or '' for linear my ($graphtype, $datafile, $title,$xlabel,$ylabel,$yrange,$yloglinear)=@_; # yrange for reads: 0.1, and for contigs: 0.9 my $graphstyle=''; if ($graphtype eq 'histogram') {$graphstyle="plot \"$datafile\" using 1:2 with boxes";} elsif ($graphtype eq 'line') {$graphstyle="plot \"$datafile\" using 1 with lines";} else {die "\n** ERROR: Invalid graphtype='$graphtype'\n\n";} my $yloglinearscale= ($yloglinear eq '') ? '' : "set $yloglinear"; # To capture any errors that are normally sent from gnuplot to stderr, could use: open3 pipe interface: # http://www.clintoneast.com/articles/perl-open3-example.php # http://hell.org.ua/Docs/oreilly/perl2/prog/ch16_03.htm # But the following should be fine, as the stderr will display when running the script anyway. # If needed a simpler way would be to sent the output to a file using eg: open (GNUPLOT, "|gnuplot > gnu_out.txt 2>&1") or die .... The read the resulting file. open (GNUPLOT, "|gnuplot") or die "\n**ERROR: Failed to open gnuplot : $!\n\n **"; print GNUPLOT <<ENDPLOT; set terminal png set output "$datafile.png" set nokey $yloglinearscale set xlabel "$xlabel" set ylabel "$ylabel" set yrange [$yrange:] set title "$title" $graphstyle ENDPLOT close(GNUPLOT); if ($? != 0) {print "\n** WARNING: GNUplot pipe returned non-zero status: '$?'\n\n";} # $? is the status returned by the last pipe close (or backtick or system operator) if (! -e "$datafile.png") {die "\n** ERROR: Failed to create '$datafile.png'**\n\n";} =pod #PNG set term png small xFFFFFF set output "$file.png" set size 1 ,1 set nokey set data style line set xlabel "frequency" font "VeraMono,10" set title "Fast Fourier Transform" font "VeraMono,10" set grid xtics ytics set xtics 100 plot "$file" using 1:2 w lines 1 =cut #WAS PREVIOUSLY AS .sh script =pod # The 'gnuplot_readshistogram_logY.sh' is: set terminal png set output "$1.png" set log y set xlabel "Read length" set ylabel "Frequency" set yrange [0.9:] set title "Histogram of read lengths" plot "$1" using 1:2 with boxes =cut } # Was previously a separate .sh file: =pod #!/bin/sh gnuplot << EOF set terminal png set output "$1.png" set xlabel "Number of contigs" set ylabel "Summed contig length in bases" set yrange [0.9:] set title "Summed contig lengths" plot "$1" using 1 with lines EOF =cut # Added function to count number of simple dinucleotide repeats: sub countRepeats { # To count the number of sequences that contain mostly repeats. # This would be faster if called a C program on the file. # Common simple repeats are listed here: http://www.bioinfo.de/isb/2005/05/0041/ # Dinucleotide # AT/TA # AC/TG # AG/TC # CG/GC # Trinucleotide # AAT/TTA # CTA/GAT # ATG/TAC # ACT/TGA # CTC/GAG # AGG/TCC # CAG/GTC # AAG/TTC # ATA/TAT # CAA/GTT # AGC/TCG # ACA/TGT # ACG/TGC # AGA/TCT # ACC/TGG # Other # Tetranucleotide # AAAT # AAAC # CACG # AACA # AATA # AAGA # TGAA # AAAG # ACAT # AATG # AGCC # Other # Pentanucleotide # AAAAC # AATTG # GCTAA # ATAAT # AAAAT # AAACA # ATATA # TTGCC # Other # I also add mono-nucleotide repeats: - ie. just all T's, or A's, etc # Just consider the dinucleotide repeats for now: my ($ATseq,$CGseq,$ACseq,$TGseq,$AGseq,$TCseq)=(0,0,0,0,0,0); my ($AAseq,$TTseq,$CCseq,$GGseq)=(0,0,0,0); foreach (@sequences) { my $seq_len=$_->[0]; my $seq=$_->[2]; # This copy might be slow, maybe should just stick with using the reference. my $mnt=0.35*$seq_len; # Mononucleotide threshold: HERE 0.35 also means 70%; 0.4 means 80% dinucleotide repeats, as really one base so 0.5 = 100% my $dnt=0.35*$seq_len; # Dinucleotide threshold: 0.35 means 70%; 0.4 means 80% dinucleotide repeats, as two bases so 0.5 = 100% my ($AT,$CG,$AC,$TG,$AG,$TC)=(0,0,0,0,0,0); my ($AA,$TT,$CC,$GG)=(0,0,0,0); # See: http://www.allinterview.com/showanswers/76719.html # AT/TA seems most common repeat so process it first to save time: while ($seq=~/AT/g) {$AT++;} if ($AT>$dnt) {$ATseq++; next;} # AT is same as TA. If has 80% AT's then won't have 80% AC etc. while ($seq=~/CG/g) {$CG++;} if ($CG>$dnt) {$CGseq++; next;} # CG is same as GC. # AC,TG while ($seq=~/AC/g) {$AC++;} if ($AC>$dnt) {$ACseq++; next;} while ($seq=~/TG/g) {$TG++;} if ($TG>$dnt) {$TGseq++; next;} # AG/TC while ($seq=~/AG/g) {$AG++;} if ($AG>$dnt) {$AGseq++; next;} while ($seq=~/TC/g) {$TC++;} if ($TC>$dnt) {$TCseq++; next;} # Added my simple mononucleotde repeat count: while ($seq=~/AA/g) {$AA++;} if ($AA>$mnt) {$AAseq++; next;} while ($seq=~/TT/g) {$TT++;} if ($TT>$mnt) {$TTseq++; next;} while ($seq=~/CC/g) {$CC++;} if ($CC>$mnt) {$CCseq++; next;} while ($seq=~/GG/g) {$GG++;} if ($GG>$mnt) {$GGseq++; next;} } my $num_seq=($#sequences+1); my $total_din_repeats_seq= $ACseq+$TGseq+$ATseq+$AGseq+$TCseq+$CGseq; my $percent_din_repeats=100*$total_din_repeats_seq/$num_seq; print STATS "\nSimple Dinucleotide repeats:\n"; printf STATS "\tNumber of %s with over 70%% dinucleotode repeats:\t%.2f %% (%d %s)\n", $type_plural, $percent_din_repeats, $total_din_repeats_seq, $type_plural; printf STATS "\tAT:\t%.2f %% (%d %s)\n", (100*$ATseq/$num_seq),$ATseq,$type_plural; printf STATS "\tCG:\t%.2f %% (%d %s)\n", (100*$CGseq/$num_seq),$CGseq,$type_plural; printf STATS "\tAC:\t%.2f %% (%d %s)\n", (100*$ACseq/$num_seq),$ACseq,$type_plural; printf STATS "\tTG:\t%.2f %% (%d %s)\n", (100*$TGseq/$num_seq),$TGseq,$type_plural; printf STATS "\tAG:\t%.2f %% (%d %s)\n", (100*$AGseq/$num_seq),$AGseq,$type_plural; printf STATS "\tTC:\t%.2f %% (%d %s)\n", (100*$TCseq/$num_seq),$TCseq,$type_plural; my $total_mono_repeats_seq= $AAseq+$TTseq+$CCseq+$GGseq; my $percent_mono_repeats=100*$total_mono_repeats_seq/$num_seq; print STATS "\nSimple mononucleotide repeats:\n"; printf STATS "\tNumber of %s with over 50%% mononucleotode repeats:\t%.2f %% (%d %s)\n", $type_plural, $percent_mono_repeats, $total_mono_repeats_seq, $type_plural; printf STATS "\tAA:\t%.2f %% (%d %s)\n", (100*$AAseq/$num_seq),$AAseq,$type_plural; printf STATS "\tTT:\t%.2f %% (%d %s)\n", (100*$TTseq/$num_seq),$TTseq,$type_plural; printf STATS "\tCC:\t%.2f %% (%d %s)\n", (100*$CCseq/$num_seq),$CCseq,$type_plural; printf STATS "\tGG:\t%.2f %% (%d %s)\n", (100*$GGseq/$num_seq),$GGseq,$type_plural; return $percent_din_repeats+$percent_mono_repeats; } sub commify_if { # If doCommify is >0 then converts output to commas. # Formats '1234567890.01' with commas as "1,234,567,890.01 # Based on: http://www.perlmonks.org/?node_id=110137 my ($number,$doCommify)=@_; if ($doCommify > 0) {$number =~ s/(\d)(?=(\d{3})+(\D|$))/$1\,/g;} return $number; } #--------------- Produce ordered list of random numbers ------------------- # This is copied from: my_random_contigs.pl sub getListOfRandomNumbers { # Use: @list=getListOfRandomNumbers(200,20); to return sorted list of 20 numbers in range from 0 to 200 inclusive. my %list2=(); my $i=0; my $MaxNumber=$_[0]; my $NumToPick=$_[1]; while ($i<$NumToPick) { my $intRand = int(rand($MaxNumber+1)); # For Zero-based perl-arrays. The +1 means this generates random integers between 0 and $MaxNumber. (See: http://perldoc.perl.org/functions/rand.html ) if ($intRand>$MaxNumber) {$intRand=$MaxNumber} # Just to be extra sure that don't exceed $MaxNumber. if ( !exists($list2{$intRand}) ) {$list2{$intRand}=1; $i++;} } #foreach my $key(keys %list2) {print "$key ";} # Sort the list of numbers: #my @SortedList2 = sort { $a <=> $b } keys(%list2); #return @SortedList2; return (sort { $a <=> $b } keys(%list2)); #print "Sorted list of ".$NumToPick." random numbers:\n"; #foreach my $num(@SortedList2) {print "$num\n";} #print "\n\n"; } sub print_sequences { # Print the sequences wrapping sequences using index array, at line length of '$fastaLineLen' characters: # Uses the global '@sequences' array. my $sequence_indexes_list=$_[0]; # This is an array reference, not the array itself. foreach my $num(@{$sequence_indexes_list}) { #print "$num (max=$#sequences)\n"; print STATS $sequences[$num]->[1]; # Prints the header, no "\n" needed after it. my $pos=0; my $seqlen=$sequences[$num]->[0]; while ($pos<$seqlen) { print STATS substr($sequences[$num]->[2],$pos,$fastaLineLen)."\n"; $pos+=$fastaLineLen; } print STATS "\n"; } } =pod # Some test runs for mono-nucleotides and dinucelotides: >FUOMOGO01AQV42DUMMYA length=339 xy=0189_0676 region=1 run=R_2009_04_23_17_54_06_ AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA >FUOMOGO01AQV42DUMMYB length=339 xy=0189_0676 region=1 run=R_2009_04_23_17_54_06_ AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA AAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAAA >FUOMOGO01AQV42DUMMYC length=339 xy=0189_0676 region=1 run=R_2009_04_23_17_54_06_ TTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTT TTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTT TTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTT TTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTTT >FUOMOGO01AQV42DUMMYD length=339 xy=0189_0676 region=1 run=R_2009_04_23_17_54_06_ GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG GGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGGG >FUOMOGO01AQV42 length=339 xy=0189_0676 region=1 run=R_2009_04_23_17_54_06_ TGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTG TGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTG TGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTG TGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTG TGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTG TGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGT >FUOMOGO01AUK0D length=214 xy=0231_0843 region=1 run=R_2009_04_23_17_54_06_ ACACACACACACACACACACACACACACACACACACACACACACACACACACACACACAC ACACACACACACACACACACACACACACACACACACACACACACACACACACACACACAC ACACACACACACACACACACACACACACACACACACACACACACACACACACACACACAC ACACACACACACACACACACACGACGACGACGAC >FUOMOGO01AUB7C length=64 xy=0228_1718 region=1 run=R_2009_04_23_17_54_06_ ATATATATATATATATATATATATATATATATATATATATATATATATATAGTACGTACG TACG >FUOMOGO01AU00B length=213 xy=0236_1097 region=1 run=R_2009_04_23_17_54_06_ ACACACACACACACACACACACACACACACACACACACACACACACACACACACACACAC ACACACACACACACACACACACACACACACACACACACACACACACACACACACACACAC ACACACACACACACACACACACACACACACACACACACACACACACACACACACACACAC ACACACACACACACACACACGACGACGACGACG >FUOMOGO01ATYRT length=169 xy=0224_0695 region=1 run=R_2009_04_23_17_54_06_ TGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTG TGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTG TGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGT >FUOMOGO01ARMLN length=400 xy=0197_2201 region=1 run=R_2009_04_23_17_54_06_ TATATATATATATATATATATATATATATATATATATATATATATATATATATATATATA TATAGTAGTAGTAGTATATATATATATATATATATATATATATATATATATATATATATA TATATATATATATATATATATATATATATATATATATATATATATATATATATATATATA TATATATATATATATATATATATATATATATATATATATATATATATATATATATATATA TATATATATATATATATATATATATATATATATATATATATATATATATATATATATATA TATATATATATATATATATATATATATATATATATATATATATATATATATATATATATA TATATATATATATATATATATATATATATATATATATATA >FUOMOGO01AVGRX length=44 xy=0241_1051 region=1 run=R_2009_04_23_17_54_06_ TATATATATATATATATATATATATATATATATATATATATATA >FUOMOGO01ASZ6K length=315 xy=0213_0922 region=1 run=R_2009_04_23_17_54_06_ TGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTG TGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTG TGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTG TGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTG TGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTGTG TGTGTGTGTGTGTGT >FUOMOGO01ARSZF length=65 xy=0199_2281 region=1 run=R_2009_04_23_17_54_06_ TATATATATATATATATATATATATATATATATATATATATATATATATATAGTACGTAC GTACG >FUOMOGO01AYV8U length=49 xy=0280_1324 region=1 run=R_2009_04_23_17_54_06_ ATATATATATATATATATATATATATATATATATATATATATATATATA >FUOMOGO01AYV9X length=40 xy=0280_1363 region=1 run=R_2009_04_23_17_54_06_ TATATATATATATATATATATATATATATATATATATATA >FUOMOGO01AUX4M length=40 xy=0235_1460 region=1 run=R_2009_04_23_17_54_06_ TATATATATATATATATATATATATATATATATATATATA >FUOMOGO01AWOTU length=54 xy=0255_0800 region=1 run=R_2009_04_23_17_54_06_ ATATATATATATATATATATATATATATATATATATATATATATATATATAGTA >FUOMOGO01A11TC length=66 xy=0316_1054 region=1 run=R_2009_04_23_17_54_06_ ATATATATATATATATATATATATATATATATATATATATATATATATATATAGTACGTA CGTACG >FUOMOGO01ASRJP length=401 xy=0210_2019 region=1 run=R_2009_04_23_17_54_06_ TATATATATATATATATATATATATATATATATATATATATATATATATATATAGTATAT AGTAGTAGTAGTATATATATATATATATATATATATATATATATATATATATATATATAT ATATATATATATATATATATATATATATATATATATATATATATATATATATATATATAT ATATATATATATATATATATATATATATATATATATATATATATATATATATATATATAT ATATATATATATATATATATATATATATATATATATATATATATATATATATATATATAT ATATATATATATATATATATATATATATATATATATATATATATATATATATATATATAT ATATATATATATATATATATATATATATATATATATATATA >FUOMOGO01AU1ZH length=67 xy=0236_2363 region=1 run=R_2009_04_23_17_54_06_ TATATATATATATATATATATATATATATATATATATATATATATATATATATAGTACGT ACGTACG =cut