1. Perl II Part III: Motifs and Loops

2. Objectives Search for motifs in DNA or Proteins Interact with users at the keyboard Write data to files Use loops Use basic regular expressions Responding to conditional tests Examining sequence data in detail

3. Conditional Tests if (1 == 1) { print “1 equals 1\n”; } if (1) { print “What does this evaluate to?\n”; } if (1 == 0) { print “1 equals 0\n”; } if (0) { print “1 evaluates to true\n”; }

4. Conditional if/else if (1 == 1) { print “1 equals 1\n\n”; } else { print “1 does not equal 1\n\n”; } unless (1 == 0) { print “1 does not equal 0\n\n”; } else { print “1 does 0?\n\n”; } Conditionals also use: ==, !=, >=,, For text: “” and ‘’ evaluate to true

5. More conditionals … #!usr/bin/perl –w #if-elseif-else $word = “MNIDDKL”; if ($word eq ‘QSTLV’) { print “QSTLV\n”; } elseif ($word eq ‘MSRQQNKISDH’) { print “MSRQQNKISDH\n”; } else { print “What is \”$word\”?\n”; exit;

6. Using Loops to Open and Read Files #!/usr/bin/perl –w $proteinFilename = “NM_012345.pep”; #open the file and catch the error unless (open(MUPPETFILE, $proteinFilename) ) { print “Could not open file $proteinFilename!\n”; exit; } #read data using a while loop, and print while ($protein = ) { print “##### Here is the next line of the file:\t”; print $protein,”\n”; } close MUPPETFILE; exit;

7. Motif finding – www.expasy.ch/prosite/ Something genuinely useful Program Flow – Reads in protein sequence from file Puts all sequence data into one string for easy searching Looks for motifs that the user types into the keyboard

8. #!/usr/bin/perl –w #searching for motifs #Ask the user for the filename of the data file print “Please type the filename of the data file: “; $proteinFilename = ; chomp $proteinFilename; #Open the file or exit open (PROTEINFILE, $proteinFilename) or die (“Error: $!”); #Read file into an array and close @protein = ; close PROTEINFILE; #Put data into a single string to make it easier to search $protein = join(‘’, @protein); $protein =~ s/[\s\t\f\r\n ]//g; Reading: ”<filename” Writing: “>filename”, discard current contents if it already exists Append: “>>filename”, open or create file for writing at end of file Update: “+<filename”, open a file for update (reading and writing) New Update: “+>filename”, create file for update is non-existent This operator will read data in until it reached the special $/ character, which is set to default as \n

9. #Ask the user for a motif, search for it, and report #if it was found. Exit if no motif was entered. do { print “Enter a motif to search for: “; $motif = ; chomp $motif; if ($protein =~ m/$motif/) { print “I found it!\n\n”; } else { print “I couldn’t find it!\n\n”; } #exit on user prompt } until ($motif =~ /^\s*$/); exit;

10. Regular Expressions Very powerful methods for matching wildcards to strings Very cryptic Perl reads =~ /n/ as =~ m/n/ The delimiter is flexible, it acccepts any nonalphanumeric nonwhitespace character (eg. #({[,.’)

11. Metasymbols SymbolMeaningSymbolMeaning \0Null Char (ASCII NULL)\NNNChar given in octal, to 377 \nNth previously captured string\aMatch the alarm character \ATrue, at the beginning of a string\bMatch the backspace char \bTrue, at word boundary\BTrue when not a word boundary \cXMatch the control char\dAny digit \DAny nondigit\eMatch the escape char \E\fFormfeed \G\nMatch the newline (NL or CR) \rMatch the return char\sMatch any whitespace \SMatch any nonwhitespace\tMatch any tab (HT) char \wMatch any ‘word’ char _AZ09\WMatch any nonword char \x{abcd}Match the char given in hex\zTrue at end of string \Z\z or before newline()Pattern brackets [ ]Either/or pattern brackets|or {n}Match only n characters.Any char but a newline *Zero or more times$Occurring at end of string ^Beginning of string{n,m}Match n to m char, inclusive ?Zero or one occurrences+One or more occurrences !~Non-match

12. Look-behind assertion (?<=value1)value2 $string = “English goodly spoken here”; $string =~ s/(?<=English )goodly/well/; (?=value1)value2 : look ahead (!=value1)value2 : not look ahead (!<=value1)value2 : not look behind

13. Backreferences Pattern == “2y 4 2 22y2y” $string =~ /(\d\w)\s+(\d)\s+(\d)\s\3\1\1/; backreferencing works within brackets from left to right

14. #!/usr/bin/perl –w #determining the frequency of nucleotides #Ask the user for the filename of the data file print “Please type the filename of the data file: “; $dnaFilename = ; ? $dnaFilename; #Open the file or exit open (DNA, $dnaFilename) or die (“Error: ?”); #Read file into an array and close @dna = ; close DNA; #Put data into a single string to make it easier to search $dna = join(‘’, @dna); $dna =~ s/[\s\t\f\r\n ]//g;

15. #Explode the $dna string into an array where it will be #easier to iterate through them and count their numbers @dna = split(‘’,$dna); #Initialize the counts $A_Number = 0; $C_Number = 0; $G_Number = 0; $T_Number = 0; $Errors = 0;

16. #Loop through the bases, examine each to determine what #each nucleotide is and increment the appropriate number foreach $base (@dna) { if ($base eq ‘A’) ++$A_Number; elseif ($base eq ‘C’) ++$C_Number; elseif ($base eq ‘G’) ++$G_Number; elseif ($base eq ‘T’) ++$T_Number; else { print “Error: I don’t recognize the base\n”; ++$Errors; } print “Base\tNumber\nA=\t$A_Number\nB=\t$B_Number\n”; print “C=\t$C_Number\nG=\t$G_Number\n\n”;

17. foreach $base (@dna) { if ($base eq ‘A’) ++$A_Number; elseif ($base eq ‘C’) ++$C_Number; elseif ($base eq ‘G’) ++$G_Number; elseif ($base eq ‘T’) ++$T_Number; else { print “Error: I don’t recognize the base\n”; ++$Errors; } foreach (@dna) { if (/A/) ++$A_Number; elseif (/C/) ++$C_Number; elseif (/G/) ++$G_Number; elseif (/T/) ++$T_Number; else { Print “Error when reading base\n”; ++$Errors; } }

18. Tricky little ifs if ($string =~ /\d{3,4}/) print “the string is 3 to four characters long\n”; = print “the string is 3 to four characters long\n” if ($string =~ /\d{3,4}/);

19. #!/usr/bin/perl –w #determining the frequency of nucleotides #Ask the user for the filename of the data file print “Please type the filename of the data file: “; $dnaFilename = ; chomp $dnaFilename; #See if the file exists then open it unless( -e $dnaFilename) { print “\”$dnaFilename\” does not exist”; exit; } open (DNA, $dnaFilename) or die (“File Error”); #Put data into a single string to make it easier to search $dna = join(‘’, @dna); $dna =~ s/[\s\t\f\r\n ]//g; Let’s do the same thing but save on some memory by not creating an array @dna = ; close DNA;

20. #Initialize the counts $A_Number = 0; $C_Number = 0; $G_Number = 0; $T_Number = 0; $Errors = 0;

21. #Loop through the bases, examine each to determine what #each nucleotide is and increment the appropriate number for ($position=0; $position<length $dna; ++$position) { $base = substr($dna, $position, 1); $_ if ($base eq ‘A’) ++$A_Number; elseif ($base eq ‘C’) ++$C_Number; elseif ($base eq ‘G’) ++$G_Number; elseif ($base eq ‘T’) ++$T_Number; else { print “Error: I don’t recognize the base\n”; ++$Errors; } print “Base\tNumber\nA=\t$A_Number\nB=\t$B_Number\n”; print “C=\t$C_Number\nG=\t$G_Number\n\n”; while($base =~ /a/ig){$a++} while($base =~ /c/ig){$c++} while($base =~ /g/ig){$g++} while($base =~ /t/ig){$t++} while($base !~ /[acgt]/ig){$e++}

22. Writing to files #All text data can be written to files $outputfile = “results.txt”; open(RESULTS, “>$ouputfile”) or die (“Error: $!”); print RESULTS “These results are overwriting everything that existed in the file results.txt\n”; Close RESULTS;

23. Command line arguments and subroutines #!/usr/bin/perl –w use strict; #Arguments collected on the command line go into a special var # called @ARGV and the program name resides in the var $0 my($title) = “$0 DNA\n\n”; unless(@ARGV) { print $title; exit; } my($input) = @ARGV[0]; print $input,”\n\n”; exit;

24. Command line arguments and subroutines #!/usr/bin/perl –w use strict; #Arguments collected on the command line go into a special var # called @ARGV and the program name resides in the var $0 my($title) = “$0 DNA\n\n”; unless(@ARGV) { print $title; exit; } my($input) = @ARGV[0]; my($subRoutineResults) = Find_Length($input); print “the length of your input is $subRoutineResults\n”; exit; sub Find_Length { my($tmp) = @_; $results = length($tmp); $return $results; }

25. Passing by value vs reference Simple routines pass everything by value However, because of the subroutine array, @_, values of arrays, hashes and scalers get flattened. Ex. my @i = (1..10); my @j = (1..23); reference_sub(@i, @j); sub { my (@i, @j) = @_; print @i.”\n@j”; }

26. my @i = (1..10); my @j = (1..23); reference_sub(\@i, \@j); #returned arrays can be referenced by @ but are global print “@i\n”; sub { my ($i, $j) = @_; print $$j[2]; push(@$i, ‘4’); }