Computational Theory MAT542 (Computational Methods in Genomics) - Part 2 & 3 -
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1 Computational Theory MAT542 (Computational Methods in Genomics) - Part 2 & 3 - Benjamin King Mount Desert Island Biological Laboratory bking@mdibl.org
2 Overview of 4 Lectures Introduction to Computation and Programming Mon. Sept. 14 Programming (Text File Processing) Wed. Sept 16 & Mon. Sept 21 Genome Sequencing and Informatics Wed. Sept 23 Homework Due on Oct. 7 th (Wed) by 2pm
3 Example Scripts and Input File go_bears.pl array_example.pl associative_array_example.pl if_loop_example.pl regular_expression_example.pl for_and_while_loop_examples.pl subsitution_and_translation_examples.pl read_file.pl ben_input_file.txt reading_and_writing_files_example.pl
4 Perl Comprehensive Perl Archive Network Active Perl (for PC, Mac OS X, Linux)
5 Homework Due by 2pm on Wednesday, Oct 7th scripts as text file attachments as well as input data files
6 Homework Assignment Write a Perl script for each of the following: 1. (10 points) Using an iterative loop and a formula, print out the following two-column array: (20 points) Print out the transcribed RNA sequence for a DNA sequence in FASTA format. The script shall read in a text file containing the input DNA sequence from a FASTA formatted sequence file. Use the GenBank record, M15131, as the input sequence. 3. (30 points) Read in a tab-delimited text file downloaded using Ensembl s BioMart that contains a listing of all transcription factors in the mouse genome, store the genome coordinates in associative arrays (using gene symbol as the key), and write an output file that contains the coordinates for all members of the HOX gene family. The list of all transcription factors can be retrieved by filtering by genes with proteins that have been annotated with the Gene Ontology molecular function term sequence-specific DNA binding transcription factor activity (GO: ). 4. (40 points) Calculate the percent GC content for each of the 36 positions in a subset of 100,000 RNAseq reads that you can download here as a FASTQ-formatted text file:
7 Programming Concepts Variables Used to store: character string integer real number Boolean value (True or False) $a = Go Bears ; $b = 25; $c = ; $d = 0; Data Structures Store collections of data in an organized fashion Common Operations Mathematical operations Testing for specific values (if / then loop) Iteration (for, while loops) Translation operations Printing messages Reading in files Writing output
8 To Run Perl Using Interactive Console 1. Type (in Command Prompt or Terminal window) perl 2. Type statements $a = 1; $b = 2; $c = $a + $b; print $c; 3. Enter CTRL-D to execute commands
9 #!/usr/bin/perl # Header # Example script # Variable declarations $a = "Go "; $b = "Black "; $c = "Bears"; go_bears.pl # Main print $a,$b,$c,"\n"; perl go_bears.pl Go Black Bears
10 Variables $a = "TAATAA"; print $a; $n = 25; $m = 100; $sum = $n + $m; Scalar Types: character string integer real number Boolean value (True or False)
11 Data Structures Store collections of data in an organized fashion Arrays ordered list of items of the same type (character, integer, = ("TAATAA", "TCATAA", "GAATAA");! print $sequences[0];! print $sequences[1];! print = (18,25,78); print $numbers[0]; print $numbers[1]; print $numbers[2];
12 Data Structures Associative Arrays list of items of the same type (character, integer, etc), but indexed by a particular character, integer, etc. %genbankids; $genbankids{"il1b"} = "M15131"; $genbankids{"hoxc8"} = "AF198989"; print $genbankids{"il1b"}; M15131 Also called hash tables Called dictionaries in Python
13
14 Sequence Alignment Program, BLAT Steps for cdna alignment: 1 Break cdna into non-overlapping n base chunks (k-mers) 2 Use index to find regions in genome similar to each k-mer 3 Find exons by looking for k-mers that align to same genome region and cdna 4 Stitch together exons
15 Sequence Alignment Program, BLAT genome: cacaattatcacgaccgc (K = 8-13 real genome) K-mers: cac aat tat cac gac cgc genome position cdna: aattctcac 3-mers: aat att ttc tct ctc tca cac hits: aat 0,3-3 cac 6,0 6 cac 6,9-3 cdna position clump: cacaattatcacgaccgc example from Jim Kent
16 Common Operations Mathematical operations Testing for specific values (if / then loop) - Regular expressions Iteration (for, while loops) Translation operations Printing messages Reading in files Writing output
17 Mathematical Operations Syntax Description Addition 10-5 Subtraction 10 * 5 Multiplication 5 / 10 Division 10**2 Exponent exp(2) log(256) Exponential function Natural log abs, atan2, cos, exp, hex, int, log, oct, rand, sin, sqrt, srand abs(-1) sqrt(256)
18 if, then loops (testing for values) $a = 1;! if ($a == 1) {! print Value is 1 ;! print hello ;! }! else {! print Value is not 1 ;! }! if ($a >= 0) {! ==!!=! >! <! >=! <=! $a!= -1! $a > 0! $b eq okay! $b ne okay!
19 if, then loops (testing for values) $a = 1;! if ($a == 1) {! print "Value is 1\n";! }! elsif ($a == 2) {! print "Value is 2\n";! }! else {! print "Value is not 1 or 2\n";! }!
20 Regular Expressions Used to match a pattern of characters Often applied in if/then loops $a = Today is Sept 11, 2013 ;! if ($a =~ /, \d+/) {! print "Found year\n";! }! if ($a =~ /, (\d+)/) {! print "Year=",$1;! }! Found year! Year=2013!
21 Regular Expressions ^ Match at beginning of string $ Match at end of string. Match any character \w Match "word" character (alphanumeric plus "_") \W Match non-word character \s Match whitespace character \S Match non-whitespace character \d Match digit character \D Match non-digit character \t Match tab \n Match newline * Match 0 or more times + Match 1 or more times? Match 1 or 0 times {n} Match exactly n times {n,} Match at least n times {n,m} Match at least n but not more than m times [ ] Match a range of characters (e.g, [A T G C] ) [0-9] [a-za-z]
22 Iteration (for, while loops) for ($i = 0; $i <= 5; $i++) {! print "i=",$i, " i**2=",$i**2, "\n";! }! i=0 i**2=0! i=1 i**2=1! i=2 i**2=4! i=3 i**2=9! i=4 i**2=16! i=5 i**2=25!
23 Iteration (for, while loops) $i = 0;! while ($i <= 5) {!!print "i=",$i, " i**2=",$i**2, "\n";!!$i = $i + 1;!!#$i++;! }! i=0 i**2=0! i=1 i**2=1! i=2 i**2=4! i=3 i**2=9! i=4 i**2=16! i=5 i**2=25!
24 Substitution and Translation Operations $sentence = "I flew to london yesterday";! $sentence =~ s/london/london/;! #$sentence =~ s/london/london/g;! print $sentence,"\n";! $sentence = "abcdefghijklmnopqrstuvwxyz";! print $sentence,"\n";! $sentence =~ tr/abc/edf/;! print $sentence,"\n";! $sentence =~ tr/[a-z]/[a-z]/;! print $sentence,"\n";! I flew to London yesterday! abcdefghijklmnopqrstuvwxyz! edfdefghijklmnopqrstuvwxyz! EDFDEFGHIJKLMNOPQRSTUVWXYZ!
25 Printing Messages $a = 24.56;! print "Value of a=",$a, "\n";! print "Value of a=$a\n";! \n = new line character \t = tab character
26 Reading Input Files Il1b Il Il10 #!/usr/bin/perl! ben_input_file.txt read_file.pl # Header! # Example script that reads in an input file! # and prints it out! # File handling! $input_fh = open(input,"<ben_input_file.txt");! # Main! while (<INPUT>) {! $line = $_;! chomp($line);! print $line, \n ;! if ($line =~ /Il12/) {! print "Found Il12\n";! }! }!
27 Reading Input Files !Il1b! !Il12! 2! !Il10! ben_input_file.txt #!/usr/bin/perl! # File handling! $input_fd = open(input,"<ben_input_file.txt");! # Main! while (<INPUT>) {!!$line = $_;!!chomp($line);!!@fields = split("\t",$line); # splits current line by! # tab characters!!$chr = $fields[0];!!$start = $fields[1];!!$symbol = $fields[2];!!print "chr=",$chr," symbol=",$symbol,"\n";! }!
28 !Il1b! !Il12! 2! !Il10! #!/usr/bin/perl! Writing Output ben_input_file.txt # File handling! $input_fd = open(input,"<ben_input_file.txt");! $output_fd = open(output,">ben_output_file.txt");! # Main! while (<INPUT>) {!!$line = $_;!!chomp($line);!!@fields = split("\t",$line); # splits current line by! # tab characters!!$chr = $fields[0];!!$start = $fields[1];!!$symbol = $fields[2];!!print OUTPUT "chr=",$chr," symbol=",$symbol,"\n";! }!
29 Using Modules
30 BioPerl
31 Install BioPerl using Active Perl s Perl Package Manager
32
33 Using Modules getlengths.pl ben_sequences.fa fasta >gi emb CAG IL1B [Homo sapiens] MAEVPKLASEMMAYYSGNEDDLFFEADGPKQMKCSFQDLDLCPLDGGIQLRISDHHYSKGFRQAASVVVA MDKLRKMLVPCPQTFQENDLSTFFPFIFEEEPIFFDTWDNEAYVHDAPVRSLNCTLRDSQQKSLVMSGPY ELKALHLQGQDMEQQVVFSMSFVQGEESNDKIPVALGLKEKNLYLSCVLKDDKPTLQLESVDPKNYPKKK MEKRFVFNKIEINNKLEFESAQFPNWYISTSQAENMPVFLGGTKGGQDITDFTMQFVSS >gi gb AAH Il1b protein [Danio rerio] MACGQYEVTIAPKNLWETDSAVYSDSDEMDCSDPLAMSYRCDMHEGIRLEMWTSQHKMKQLVNVIIALNR MKHIKPQSTEFGEKEVLDMLMANVIQEREVNVVDSVPSYTKTKNVLQCTICDQYKKSLVRSGGSPHLQAV TLRAGSSDLKVRFSMSTYASPSAPATSAQPVCLGISKSNLYLACSPAEGSAPHLVLKEISGSLETIKAGD PNGYDQLLFFRKETGSSINTFESVKCPGWFISTAYEDSQMVEMDRKDTERIINFELQDKVRI >gi ref NP_ interleukin 1, beta [Rattus norvegicus] MATVPELNCEIAAFDSEENDLFFEADRPQKIKDCFQALDLGCPDESIQLQISQQHLDKSFRKAVSLIVAV EKLWQLPMSCPWSFQDEDPSTFFSFIFEEEPVLCDSWDDDDLLVCDVPIRQLHCRLRDEQQKCLVLSDPC ELKALHLNGQNISQQVVFSMSFVQGETSNDKIPVALGLKGLNLYLSCVMKDGTPTLQLESVDPKQYPKKK MEKRFVFNKIEVKTKVEFESAQFPNWYISTSQAEHRPVFLGNSNGRDIVDFTMEPVSS
34 Using Modules getlengths.pl ben_sequences.fa fasta # first, bring in the SeqIO module use Bio::SeqIO; # usage statement if one or both arguments are missing. my $usage = "getlengths.pl file format\n"; my $file = shift or die $usage; my $format = shift or die $usage; # create a SeqID object that will bring in the contents of the input file my $inseq = Bio::SeqIO->new(-file => "<$file", -format => $format ); while (my $seq = $inseq->next_seq) { print $seq->length,"\n"; } exit;
35 Homework Due by 2pm on Wednesday, Oct. 1 st scripts as text file attachments as well as input data files bking@mdibl.org
36 Homework Assignment Write a Perl script for each of the following: 1. (10 points) Using an iterative loop and a formula, print out the following two-column array: (20 points) Print out the transcribed RNA sequence for a DNA sequence in FASTA format. The script shall read in a text file containing the input DNA sequence from a FASTA formatted sequence file. Use the GenBank record, M15131, as the input sequence. 3. (30 points) Read in a tab-delimited text file downloaded using Ensembl s BioMart that contains a listing of all transcription factors in the mouse genome, store the genome coordinates in associative arrays (using gene symbol as the key), and write an output file that contains the coordinates for all members of the HOX gene family. The list of all transcription factors can be retrieved by filtering by genes with proteins that have been annotated with the Gene Ontology molecular function term sequence-specific DNA binding transcription factor activity (GO: ). 4. (40 points) Calculate the percent GC content for each of the 36 positions in a subset of 100,000 RNAseq reads that you can download here as a FASTQ-formatted text file:
37 Retrieve a Sequence in FASTA Format
38 Retrieve a Sequence in FASTA Format
39 Retrieve a Sequence in FASTA Format
40 Retrieve a Sequence in FASTA Format
41 Gene Ontology Uses$terms$to$describe$gene$products:$ $Biological$Process$ $Molecular$Func8on$ $Cellular$Component$ Given$term$may$have$mul8ple$parent$nodes$(DAG$=$directed$acyclic$graph)$
42 Obtain List of All Human Genes Annotated To Be Involved in Signal Transduction Using Ensembl s BioMart
43 Obtain List of All Human Genes Annotated To Be Involved in Signal Transduction Using Ensembl s BioMart
44 Obtain List of All Human Genes Annotated To Be Involved in Signal Transduction Using Ensembl s BioMart
45 Obtain List of All Human Genes Annotated To Be Involved in Signal Transduction Using Ensembl s BioMart
46 Obtain List of All Human Genes Annotated To Be Involved in Signal Transduction Using Ensembl s BioMart
47 Obtain List of All Human Genes Annotated To Be Involved in Signal Transduction Using Ensembl s BioMart Gene Ontology Biological Process Term Name signal transduction GO:
48 Obtain List of All Human Genes Annotated To Be Involved in Signal Transduction Using Ensembl s BioMart
49 Obtain List of All Human Genes Annotated To Be Involved in Signal Transduction Using Ensembl s BioMart
50 Obtain List of All Human Genes Annotated To Be Involved in Signal Transduction Using Ensembl s BioMart
51 Obtain List of All Human Genes Annotated To Be Involved in Signal Transduction Using Ensembl s BioMart
52 Obtain List of All Human Genes Annotated To Be Involved in Signal Transduction Using Ensembl s BioMart
53 Obtain List of All Human Genes Annotated To Be Involved in Signal Transduction Using Ensembl s BioMart
54 Obtain List of All Human Genes Annotated To Be Involved in Signal Transduction Using Ensembl s BioMart
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