Download Three main topics for this Intro lecture:

Bioinformatics
生物信息学理论和实践
唐继军
[email protected]
13928761660
Exercise 1
•
•
•
•
Ask for a protein file in fasta format
Ask for an amino acid
Count the frequency of that amino acid
TKFHSNAHFYDCWRMLQYQLDMRCMRAISTF
SPHCGMEHMPDQTHNQGEMCKPRMWQVS
MNQSCNHTPPFRKTYVEWDYMAKALIAPYTL
GWLASTCFIW
Exercise 2
•
•
•
•
•
Ask for an RNA file in fasta format
Convert it to RNA
Ask for a codon
Count the frequency of that codon
TCGTACTTAGAAATGAGGGTCCGCTTTTGCCC
ACGCACCTGATCGCTCCTCGTTTGCTTTTAAG
AACCGGACGAACCACAGAGCATAAGGAGAA
CCTCTAGCTGCTTTACAAAGTACTGGTTCCCT
TTCCAGCGGGATGCTTTATCTAAACGCAATGA
Subroutine
• Some code needs to be reused
• A good way to organize code
• Called "function" in some languages
• Name
• Return
• Parameters (@_)
sub codon2aa {
my($codon) = @_;
if ( $codon =~ /GC./i)
{ return 'A' }
Alanine
elsif ( $codon =~ /TG[TC]/i)
{ return 'C' }
Cysteine
elsif ( $codon =~ /GA[TC]/i)
{ return 'D' }
Aspartic Acid
elsif ( $codon =~ /GA[AG]/i)
{ return 'E' }
Glutamic Acid
elsif ( $codon =~ /TT[TC]/i)
{ return 'F' }
Phenylalanine
elsif ( $codon =~ /GG./i)
{ return 'G' }
Glycine
elsif ( $codon =~ /CA[TC]/i)
{ return 'H' }
Histidine
elsif ( $codon =~ /AT[TCA]/i)
{ return 'I' }
Isoleucine
elsif ( $codon =~ /AA[AG]/i)
{ return 'K' }
Lysine
elsif ( $codon =~ /TT[AG]|CT./i) { return 'L' }
Leucine
elsif ( $codon =~ /ATG/i)
{ return 'M' }
Methionine
elsif ( $codon =~ /AA[TC]/i)
{ return 'N' }
Asparagine
elsif ( $codon =~ /CC./i)
{ return 'P' }
Proline
elsif ( $codon =~ /CA[AG]/i)
{ return 'Q' }
Glutamine
elsif ( $codon =~ /CG.|AG[AG]/i) { return 'R' }
Arginine
elsif ( $codon =~ /TC.|AG[TC]/i) { return 'S' }
Serine
elsif ( $codon =~ /AC./i)
{ return 'T' }
Threonine
elsif ( $codon =~ /GT./i)
{ return 'V' }
Valine
elsif ( $codon =~ /TGG/i)
{ return 'W' }
Tryptophan
elsif ( $codon =~ /TA[TC]/i)
{ return 'Y' }
Tyrosine
elsif ( $codon =~ /TA[AG]|TGA/i) { return '_' }
Stop
else {print STDERR "Bad codon \"$codon\"!!\n"; exit; }
}
!/usr/bin/perl –w
print "Please type the filename: ";
$dna_filename = <STDIN>; chomp $dna_filename;
open(DNAFILE, $dna_filename);
$name = <DNAFILE>;@DNA = <DNAFILE>;close DNAFILE;
$DNA = join( '', @DNA);$DNA =~ s/\s//g;
print "First
print "Second
print "Third
", dna2peptide($DNA), "\n";
", dna2peptide(substr($DNA, 1)), "\n";
", dna2peptide(substr($DNA, 2)), "\n";
$DNA = reverse
print "Fourth
print "Fifth
print "Sixth
$DNA;
", dna2peptide($DNA), "\n";
", dna2peptide(substr($DNA, 1)), "\n";
", dna2peptide(substr($DNA, 2)), "\n";
sub dna2peptide {
my ($dna) = @_;
my $protein = "";
for(my $i=0; $i < (length($dna) - 2) ; $i += 3) {
$codon = substr($dna,$i,3);
$protein .= codon2aa($codon);
}
return $protein;
}
sub codon2aa {
...
}
Modules
• A Perl Module is a self-contained pieceof
[Perl] code that can be used by a Perl
program later
• Like a library
• End with extension .pm
• Needs a 1 at the end
Bio.pm
sub codon2aa {
....
....
}
sub dna2peptide {
....
....
}
1
!/usr/bin/perl -w
use Bio;
print "Please type the filename: ";
$dna_filename = <STDIN>; chomp $dna_filename;
open(DNAFILE, $dna_filename);
$name = <DNAFILE>;@DNA = <DNAFILE>;close DNAFILE;
$DNA = join( '', @DNA);$DNA =~ s/\s//g;
print "First
print "Second
print "Third
", dna2peptide($DNA), "\n";
", dna2peptide(substr($DNA, 1)), "\n";
", dna2peptide(substr($DNA, 2)), "\n";
$DNA = reverse $DNA;
$DNA =~ tr/ACGTacgt/TGCAtgca/;
print "Fourth
print "Fifth
print "Sixth
", dna2peptide($DNA), "\n";
", dna2peptide(substr($DNA, 1)), "\n";
", dna2peptide(substr($DNA, 2)), "\n";
Bio.pm
sub codon2aa {
....
....
}
sub dna2peptide {
....
....
}
sub fasta_read {
print "Please type the filename: ";
my $dna_filename = <STDIN>; chomp $dna_filename;
unless (open(DNAFILE, $dna_filename)) {
print "Cannot open file ", $dna_filename, "\n";
}
$name = <DNAFILE>;@DNA = <DNAFILE>;close DNAFILE;
$DNA = join( '', @DNA);$DNA =~ s/\s//g;
return $DNA;
}
1
!/usr/bin/perl -w
use Bio;
$DNA = fasta_read();
print "First
print "Second
print "Third
", dna2peptide($DNA), "\n";
", dna2peptide(substr($DNA, 1)), "\n";
", dna2peptide(substr($DNA, 2)), "\n";
$DNA = reverse $DNA;
$DNA =~ tr/ACGTacgt/TGCAtgca/;
print "Fourth
print "Fifth
print "Sixth
", dna2peptide($DNA), "\n";
", dna2peptide(substr($DNA, 1)), "\n";
", dna2peptide(substr($DNA, 2)), "\n";
Scope
• my provides lexical scoping; a variable
declared with my is visible only within the
block in which it is declared.
• Blocks of code are hunks within curly
braces {}; files are blocks.
• Use use vars qw([list of var names]) or our
([var_names]) to create package globals.
!/usr/bin/perl -w
use Bio;
use strict;
use warnings;
$DNA = fasta_read();
print "First
print "Second
print "Third
", dna2peptide($DNA), "\n";
", dna2peptide(substr($DNA, 1)), "\n";
", dna2peptide(substr($DNA, 2)), "\n";
$DNA = reverse $DNA;
$DNA =~ tr/ACGTacgt/TGCAtgca/;
print "Fourth
print "Fifth
print "Sixth
", dna2peptide($DNA), "\n";
", dna2peptide(substr($DNA, 1)), "\n";
", dna2peptide(substr($DNA, 2)), "\n";
Variable "$DNA" is not imported at frame2.pl line 6.
Variable "$DNA" is not imported at frame2.pl line 8.
Variable "$DNA" is not imported at frame2.pl line 9.
Variable "$DNA" is not imported at frame2.pl line 10.
Variable "$DNA" is not imported at frame2.pl line 12.
Variable "$DNA" is not imported at frame2.pl line 12.
Variable "$DNA" is not imported at frame2.pl line 13.
Variable "$DNA" is not imported at frame2.pl line 14.
Variable "$DNA" is not imported at frame2.pl line 15.
Global symbol "$DNA" requires explicit package name at frame2.pl
Global symbol "$DNA" requires explicit package name at frame2.pl
Global symbol "$DNA" requires explicit package name at frame2.pl
Global symbol "$DNA" requires explicit package name at frame2.pl
Global symbol "$DNA" requires explicit package name at frame2.pl
Global symbol "$DNA" requires explicit package name at frame2.pl
Global symbol "$DNA" requires explicit package name at frame2.pl
Global symbol "$DNA" requires explicit package name at frame2.pl
Global symbol "$DNA" requires explicit package name at frame2.pl
Execution of frame2.pl aborted due to compilation errors.
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!/usr/bin/perl -w
use Bio;
use strict;
use warnings;
my $DNA = fasta_read();
print "First
print "Second
print "Third
", dna2peptide($DNA), "\n";
", dna2peptide(substr($DNA, 1)), "\n";
", dna2peptide(substr($DNA, 2)), "\n";
$DNA = reverse $DNA;
$DNA =~ tr/ACGTacgt/TGCAtgca/;
print "Fourth
print "Fifth
print "Sixth
", dna2peptide($DNA), "\n";
", dna2peptide(substr($DNA, 1)), "\n";
", dna2peptide(substr($DNA, 2)), "\n";
my $x = 10;
for (my $x = 0; $x < 5; $x++) {
Scope();
print $x, "\n";
}
print $x, "\n";
sub Scope {
my $x = 0;
}
sub get_file_data {
my($filename) = @_;
use strict;
use warnings;
# Initialize variables
my @filedata = ( );
unless( open(GET_FILE_DATA, $filename) ) {
print STDERR "Cannot open file \"$filename\"\n\n";
exit;
}
@filedata = <GET_FILE_DATA>;
close GET_FILE_DATA;
return @filedata;
}
sub extract_sequence_from_fasta_data {
my(@fasta_file_data) = @_;
my $sequence = '';
foreach my $line (@fasta_file_data) {
if ($line =~ /^\s*$/) {
next;
} elsif($line =~ /^\s*#/) {
next;
} elsif($line =~ /^>/) {
next;
} else {
$sequence .= $line;
}
}
# remove non-sequence data (in this case, whitespace) from $sequence string
$sequence =~ s/\s//g;
return $sequence;
}
Molecular Scissors
Molecular Cell Biology, 4th edition
Discovering Restriction Enzymes
• HindII - first restriction enzyme – was
discovered accidentally in 1970 while
studying how the bacterium Haemophilus
influenzae takes up DNA from the virus
• Recognizes and cuts DNA at sequences:
•
•
GTGCAC
GTTAAC
Recognition Sites of Restriction Enzymes
Molecular Cell Biology, 4th edition
Uses of Restriction Enzymes
• Recombinant DNA technology
• Cloning
• cDNA/genomic library construction
• DNA mapping
Restriction Enzyme Database
• http://rebase.neb.com/rebase/rebase.html
http://rebase.neb.com/rebase/rebase.files.html
R
Y
M
K
S
W
B
D
H
V
N
=
=
=
=
=
=
=
=
=
=
=
G or A
C or T
A or C
G or T
G or C
A or T
not A (C or
not C (A or
not G (A or
not T (A or
A or C or G
G or
G or
C or
C or
or T
T)
T)
T)
G)
sub IUB_to_regexp {
my($iub) = @_;
my $regular_expression = ‘’;
my %iub2character_class = (
A
C
G
T
R
Y
M
K
S
W
B
D
H
V
N
=>
=>
=>
=>
=>
=>
=>
=>
=>
=>
=>
=>
=>
=>
=>
'A',
'C',
'G',
'T',
'[GA]',
'[CT]',
'[AC]',
'[GT]',
'[GC]',
'[AT]',
'[CGT]',
'[AGT]',
'[ACT]',
'[ACG]',
'[ACGT]',
);
$iub =~ s/\^//g;
for ( my $i = 0 ; $i < length($iub) ; ++$i ) {
$regular_expression .= $iub2character_class{substr($iub, $i, 1)};
}
return $regular_expression;
}
Hash
• Initialize: my %hash = ();
• Add key/value pair: $hash{$key}
• Add more keys:
= $value;
• %hash = ( 'key1', 'value1', 'key2', 'value2 );
• %hash = ( key1 => 'value1', key2 => 'value2', );
• Delete: delete
$hash{$key};
while ( my ($key, $value) = each(%hash) )
{
print "$key => $value\n";
}
for my $key ( keys %hash ) {
my $value = $hash{$key};
print "$key => $value\n";
}
sub codon2aa {
my($codon) = @_;
$codon = uc $codon;
my %genetic_code = (
'TCA' =>
'TCC' =>
'TCG' =>
'TCT' =>
'TTC' =>
'TTT' =>
'TTA' =>
'TTG' =>
#Many more
);
'S',
'S',
'S',
'S',
'F',
'F',
'L',
'L',
#
#
#
#
#
#
#
#
Serine
Serine
Serine
Serine
Phenylalanine
Phenylalanine
Leucine
Leucine
if(exists $genetic_code{$codon}) {
return $genetic_code{$codon};
}else{
print STDERR "Bad codon \"$codon\"!!\n";
exit;
}
}
sub parseREBASE {
my($rebasefile) = @_;
my @rebasefile = ( ); my %rebase_hash = (
); my $name; my $site;
my $regexp;
open($rebase_filehandle, $rebasefile) or die "Cannot open file\n";
while(<$rebase_filehandle>) {
# Discard header lines
( 1 .. /Rich Roberts/ ) and next;
# Discard blank lines
/^\s*$/ and next;
# Split the two (or three if includes parenthesized name) fields
my @fields = split( " ", $_);
$name = shift @fields;
$site = pop @fields;
# Translate the recognition sites to regular expressions
$regexp = IUB_to_regexp($site);
# Store the data into the hash
$rebase_hash{$name} = "$site $regexp";
}
# Return the hash containing the reformatted REBASE data
return %rebase_hash;
}
Range
• ( 1 .. /Rich Roberts/ ) and next
• from first line till some line containing Rich Roberts
• If that is true, it will check the statement after "and"
• If that is not true, it will not check the statement after
"and"
• open(…) or die
• If can open, the statement is already true, no need to
check the statement after "or"
• If cannot open, the statement is false, need to check
the statement after "or" to see if it can be true
@fred = (1,2,3);
@barney = @fred;
@huh = 1;
@fred = qw(one two);
@barney = (4,5,@fred,6,7);
@barney = (8,@barney);
($a,$b,$c) = (1,2,3);
@fred = (@barney = (2,3,4));
@fred = @barney = (2,3,4);
@fred = (1,2,3);
$fred[3] = "hi";
$fred[6] = "ho"; # @fred is now (1,2,3,"hi",undef,undef,"ho")
Array operators
• push and pop (right-most element)
• @mylist = (1,2,3); push(@mylist,4,5,6);
• $oldvalue = pop(@mylist);
• shift and unshift (left-most element)
• @fred = (5,6,7); unshift(@fred,2,3,4);
• $x = shift(@fred);
• reverse: @a = (7,8,9); @b = reverse(@a);
• sort: @a = (7,9,9); @b = sort(@a);
sub match_positions {
my($regexp, $sequence) = @_;
use BeginPerlBioinfo;
my @positions = (
);
while ( $sequence =~ /$regexp/ig ) {
push ( @positions, pos($sequence) - length($&) + 1);
}
return @positions;
}
use BeginPerlBioinfo;
my %rebase_hash = ( ); my @file_data = (
$recognition_site = '';
my $regexp = ''; my @locations = ( );
); my $query = ''; my $dna = ''; my
@file_data = get_file_data("sample.dna");
$dna = extract_sequence_from_fasta_data(@file_data);
%rebase_hash = parseREBASE('bionet');
do {
print "Search for what restriction site for (or quit)?: ";
$query = <STDIN>;
chomp $query;
if ($query =~ /^\s*$/ ) { exit;
}
if ( exists $rebase_hash{$query} ) {
($recognition_site, $regexp) = split ( " ", $rebase_hash{$query});
@locations = match_positions($regexp, $dna);
if (@locations) {
print "Searching for $query $recognition_site $regexp\n";
print "Restriction site for $query at :", join(" ", @locations), "\n";
} else {
print "A restriction enzyme $query is not in the DNA:\n";
}
}
} until ( $query =~ /quit/ );
exit;