binf634 fall015 lecture 11 binf 634 bioinformatics programming instructor: jeff solka ph.d. phone:...
TRANSCRIPT
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BINF 634 Bioinformatics Programming Instructor: Jeff Solka Ph.D. Phone: 540-809-9799 Email: [email protected] Office Hours: By appointment Required texts:
Beginning Perl for Bioinformatics by Tisdall and Waliszewski Programming Perl (3rd Edition) by Wall, Christiansen and
Orwant Course Meeting Place: Ocaquan Prince William Rm. 304B Course Meeting Times: M: 4:30 pm – 7:10 pm Course webpage
http://binf.gmu.edu/jsolka/fall15/binf634/Fall_2015BINF_634_Syllabus_rev1.html
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Acknowledgements
Some of the material used in this course was previously developed by
John Grefenstette John Kopecky
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Experimental Biology Computational Biology and Bioinformatics
Database
Problem Statement Simulation Results
Problem Statement Experiment Results
Analysis Tools
SIMS
LIMS
Computational Biology
Experimental BiologyRick Stevens
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Bioinformatics Programming Tasks Manage large experimental data sets
Sequence data Microarray data (gene expression) Mass spec data (proteomics) Genotype project data (HapMap) Clinical data
Build tools for Knowledge Discovery Find motifs in sequence data Data clustering Visualization
Build analysis pipelines Glue several analysis steps together into a single automated
process "Munge" data: Take data from one application or database and
format it for input to another application of database
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Objectives Programming skills
Problem solving and Debugging Reading and Writing Documentation Data Munging: Data filtering and transformation Pattern matching and data mining Visualization and web presentation Object-oriented programming
Bioinformatics skills Biological sequence analysis Interacting with biological databases Using Bioperl
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Background and Prerequisites Molecular Biology
BIOL 482 or similar course Recombinant DNA - Watson, Gilman, Witlowski,
Zoller http://www.amazon.com/Recombinant-DNA-Genes-Genomes-
Course/dp/0716728664/ref=dp_ob_title_bk Online Tutorials
http://www.biology-online.org/1/5_DNA.htm
Computer Science IT 108, CS 112 or similar Previous programming experience
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Course Policies Programming assignments (50%)
3-4 graded programming assignments Exams: Midterm (20%) and Final (20%)
May include both closed-book section and open-book programming problems
In-class Quizzes (10%) Weekly homework assignments
All HW assignments must be submitted to me via Blackboard by the beginning of the next class. HW assignments will not be graded individually, but you may be called upon to discuss your work during the next class. Therefore, late assignments will not be accepted.
Grading will be on the following scale. 93-100 (A), 90-92 (A-), 87-89 (B+), 83-86 (B), 80-82(B-), below 80 (C). Student averages will be rounded to the closest integer to determine final letter grades.
Keep an eye on the webpage http://binf.gmu.edu/jsolka/fall15/binf634/
Fall_2015BINF_634_Syllabus_rev1.html
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Honor Code Policies I take honor code violations very seriously.
Programming assignments must be your work. Each assignment will specify whether you may use code from other sources. Any material you take from another source must be acknowledged within the program documentation. You must read and understand the honor code handout. Violations of the honor code WILL be referred to the Honor Council.
All students must adhere to the GMU Honor Code: See:
http://www.gmu.edu/academics/catalog/9798/honorcod.html
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PragmaticsAssignments and Announcement
Will be posted on course wepage; check daily Class email will be sent to your email address from Patriot Web
Accounts You should have an account on the server binf.gmu.edu Systems administrator: Chris Ryan, [email protected]
Accessing perl: Method 1a:Login from Rooms 304B or 320 Method 1b:Login from off-campus using ssh
Go to ftp://ftp.ssh.com/pub/ssh/ for academic Windows client Alternatively go to
http://www.chiark.greenend.org.uk/~sgtatham/putty/ Method 2:Install perl on your own computer -- see textbooks and
backup slide materials
Pragmatics Unix
This class will focus on using the Unix operating system We will be using Mac OS X (at least in the classroom)
There are numerous UNIX tutorials http://www.unixtools.com/tutorials.html
Text Editors Perl program are stored in plain text files I recommend emacs or vim for a Unix text editor (see links for windows
support) http://www.claremontmckenna.edu/math/ALee/emacs/emacs.html http://www.vim.org
If you are interested in an integrated development environment I recommend Eclipse (see backup slides)
www.eclipse.org There is a tutorials for each online
http://www.gnu.org/software/emacs/tour/ http://www.yolinux.com/TUTORIALS/LinuxTutorialAdvanced_vi.html
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Review: Molecular Biology Life evolved from common
origin about 3.5 billion years ago
All life shares similar biochemistry
Proteins: active elements Nucleic acids:
informational elements
Molecular Biology: the study of structure and function of proteins and nucleic acids
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Proteins Functions:
Structural proteins Enzymes Transport Antibody defense
Structure: Chains of amino acids Typical size ~300 residues Range from about 100 to
over 5000 residues
N.B. – A residue is one of the 20 building blocks of proteins also called an amino acid.
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DNA Double stranded Four bases: adenine (A),
guanine (G), cytosine (C) and thymine (T)
A and G are purines C and T are pyrimidines A always paired with T
(complementary) C always paired with G
(complementary)=> Watson-Crick base pairs
(bp) DNA may consist of
hundreds of millions bp A short sequence (<100) is
called an oligonucleotide
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RNA:
single stranded
uses U (uracil) instead of T
less stable than DNA
also used in functional molecules (e.g. rRNA, tRNA)
rRNA = ribosomal RNA
tRNA = transfer RNA
important regulatory functions (siRNA)
siRNA = small interferring RNA
•introns are not translated
•exons are translated
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Translation
Translation involves mRNA and ribosomes
Ribosomes made of protein and ribosomal RNA (rRNA)
Transfer RNA (tRNA) make connection between specific codons in mRNA and amino acids
As tRNA binds to the next codon in mRNA, its amino acid is bound to the last amino acid in the protein chain
When a STOP codon is encountered, the ribosome releases the mRNA and synthesis ends
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DNA Structure DNA contains:
Genes "a locatable region of genomic sequence,
corresponding to a unit of inheritance, which is associated with regulatory regions, transcribed regions, and or other
functional sequence regions ".[1] Promoters
“a promoter is a region of DNA that facilitates the transcription of a particular gene”
Non-coding regions DNA which does not contain instructions
for making proteins
Reading frames An open reading frames
(ORF): a contiguous sequence of DNA starting at a start codon and ending at a STOP codon
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Shotgun DNA Sequencing
More discussions can be found herehttp://en.wikipedia.org/wiki/Shotgun_sequencing
>gi|40457238|HIV-1 isolate 97KE128 from Kenya gag gene, partial cdsCTTTTGAATGCATGGGTAAAAGTAATAGAAGAAAGAGGTTTCAGTCCAGAAGTAATACCCATGTTCTCAGCATTATCAGAAGGAGCCACCCCACAAGATTTAAATACGATGCTGAACATAGTGGGGGGACACCAGGCAGCTATGCAAATGCTAAAGGATACCATCAATGAGGAAGCTGCAGAATGGGACAGGTTACATCCAGTACATGCAGGGCCTATTCCGCCAGGCCAGATGAGAGAACCAAGGGGAAGTGACATAGCAGGAACTACTAGTACCCCTCAAGAACAAGTAGGATGGATGACAAACAATCCACCTATCCCAGTGGGAGACATCTATAAAAGATGGATCATCCTGGGCTTAAATAAAATAGTAAGAATGTATAGCCCTGTTAGCATTTTGGACATAAAACAAGGGCCAAAAGAACCCTTTAGAGACTATGTAGATAGGTTCTTTAAAACTCTCAGAGCCGAACAAGCTT
>gi|40457236| HIV-1 isolate 97KE127 from Kenya gag gene, partial cdsTTGAATGCATGGGTGAAAGTAATAGAAGAAAAGGCTTTCAGCCCAGAAGTAATACCCATGTTCTCAGCATTATCAGAAGGAGCCACCCCACAAGATTTAAATATGATGCTGAATATAGTGGGGGGACACCAGGCAGCTATGCAAATGTTAAAAGATACCATCAATGAGGAAGCTGCAGAATGGGACAGGTTACATCCAATACATGCAGGGCCTATTCCACCAGGCCAAATGAGAGAACCAAGGGGAAGTGACATAGCAGGAACTACTAGTACCCCTCAAGAGCAAATAGGATGGATGACAAGCAACCCACCTATCCCAGTGGGAGACATCTATAAAAGATGGATAATCCTGGGATTAAATAAAATAGTAAGAATGTATAGCCCTGTTAGCATTTTGGACATAAAACAAGGGCCAAAAGAACCTTTCAGAGACTATGTAGATAGGTTTTTTAAAACTCTCAGAGCCGAACAAGCTT
>gi|40457234| HIV-1 isolate 97KE126 from Kenya gag gene, partial cdsCCTTTGAATGCATGGGTGAAAGTAATAGAAGAAAAGGCTTTCAGCCCAGAAGTAATACCCATGTTTTCAGCATTATCAGAAGGAGCCACCCCACAAGATTTAAATATGATGCTGAACATAGTGGGGGGGCACCAGGCAGCTATGCAAATGTTAAAAGATACCATCAATGAGGAAGCTGCAGAATGGGACAGGCTACATCCAGCACAGGCAGGGCCTATTGCACCAGGCCAGATAAGAGAACCAAGGGGAAGTGATATAGCAGGAACTACTAGTACCCCTCAAGAACAAATAGCATGGATGACAGGCAACCCGCCTATCCCAGTGGGAGACATCTATAAAAGATGGATAATCCTGGGATTAAATAAAATAGTAAGAATGTATAGCCCTGTTAGCATTTTGGATATAAAACAAGGGCCAAAAGAACCATTCAGAGACTATGTAGACAGGTTCTTTAAAACTCTCAGAGCCGAACAAGCTT
Sequence Files -- FASTA Format
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GenBank RecordLOCUS AK091721 2234 bp mRNA linear PRI 20-JAN-2006DEFINITION Homo sapiens cDNA FLJ34402 fis, clone HCHON2001505.ACCESSION AK091721VERSION AK091721.1 GI:21750158KEYWORDS oligo capping; fis (full insert sequence).SOURCE Homo sapiens (human) ORGANISM Homo sapiens Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Euteleostomi; Mammalia; Eutheria; Euarchontoglires; Primates; Catarrhini; Hominidae; Homo. TITLE Complete sequencing and characterization of 21,243 full-length human cDNAs JOURNAL Nat. Genet. 36 (1), 40-45 (2004)FEATURES Location/Qualifiers source 1..2234 /organism="Homo sapiens" /mol_type="mRNA" CDS 529..1995 /note="unnamed protein product" /codon_start=1 /protein_id="BAC03731.1" /db_xref="GI:21750159" /translation="MVAERSPARSPGSWLFPGLWLLVLSGPGGLLRAQEQPSCRRAFD
... RLDALWALLRRQYDRVSLMRPQEGDEGRCINFSRVPSQ"ORIGIN 1 gttttcggag tgcggaggga gttggggccg ccggaggaga agagtctcca ctcctagttt 61 gttctgccgt cgccgcgtcc cagggacccc ttgtcccgaa gcgcacggca gcggggggaa
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Why Perl? Widely used in Bioinformatics
Bioperl http://www.bioperl.org/wiki/Main_Page
Ease of Programming Excellent pattern matching features Good for gluing other programs together Easy to learn (enough to get started)
Rapid Prototyping Few lines of code needed for many problems One-liners
Portability Runs on Unix, Windows, Macs
Open Source Culture Many sources of help ( try: %perldoc perldoc)
%perldoc –f print http://perldoc.perl.org/index-tutorials.html Many sources of useful modules ( http://www.cpan.org/ )
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VariablesThe types of Perl variables are indicated by the initial symbol:
$var stores a scalar (a single string or number)$x = 10;$s = "ATTGCGT";$x = 3.1417;
@var stores an array (a list of values)@a = (10, 20, 30);@a = (100, $x, "Jones", $s);print "@a\n"; # prints "100 3.1417 Jones ATTGCGT"
%var stores a hash (associative array)%ages = { John => 30, Mary => 22, Lakshmi => 27 };print $age{"Mary"}, "\n"; # prints 22
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Declaring Variables
use strict; Putting use strict; at the top of your programs will tell perl to
slap your hands with a fatal error whenever you break certain rules.
Requires us to declare all variables Avoids creating variable by typos
variables may be declaring using my, our or local for now, we only need to use my:
my $a; # value of $a is undefmy ($a, $b, $c); # $a, $b, $c are all undefmy @array; # value of @array is ()
Can combine declaration and initialization:my @array = qw/A list of words/;
my $a = "A string";
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How Things Can Go Wrong
http://www.perlmonks.org/?node_id=269642
We will come back and examine this after we have discussed references.
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Scalar and List Context All operations in Perl are evaluated in either scalar or list
context, and may behave differently depending on context
@array = ('one', 'two', 'three');$a = @array; # scalar context for assignment, return sizeprint $a; # prints 3
($a) = @array; # list context for assignmentprint $a; # prints 'one'
($a, $b) = @array;print "$a, $b"; # prints 'one, two'($a, $b, $c, $d) = @array; # $d is undefined
In computer science a list is an ordered collection of values
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String Operations Ways to concatenate strings
$DNA1 = "ATG";$DNA2 = "CCC";$DNA3 = $DNA1 . $DNA2; # concatenation operator$DNA3 = "$DNA1$DNA2"; # string interpolationprint "$DNA3"; # prints ATGCCC
$DNA3 = '$DNA1$DNA2'; # no string interpolationprint "$DNA3"; # prints $DNA1$DNA2
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ArraysAn array stores an ordered list of scalars:
@gene_array = (‘EGF1’, ‘TFEC’, ‘CFTR’, ‘LOC1691’);print “@gene_array\n”;
Output:EGF1 TFEC CFTR LOC1691
# there’s more than one way to do it (see previous slide on declaring variables)
@gene_array = qw/EGF1 TFEC CFTR LOC1691/;
http://www.perlmeme.org/howtos/perlfunc/qw_function.html
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Arrays
An array stores an ordered list of scalars:@a = (‘one’, ‘two’, ‘three’, ‘four’);
The array is indexed by integers starting with 0:
print “$a[1] $a[0] $a[3]\n”;
prints:
two one four
Notice: $a[i] is a scalar since we used the $ method of addressing the variable
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Unix Commands I cat --- for creating and displaying short files chmod --- change permissions cd --- change directory cp --- for copying files date --- display date echo --- echo argument ftp --- connect to a remote machine to download or upload files grep --- search file head --- display first part of file ls --- see what files you have lpr --- standard print command more --- use to read files mkdir --- create directory mv --- for moving and renaming files
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Unix Commands II pwd --- find out what directory you are in rm --- remove a file rmdir --- remove directory setenv --- set an environment variable sort --- sort file tail --- display last part of file tar --- create an archive, add or extract files ssh --- log in to another machine wc --- count characters, words, lines
This site has a nice reference card http://www.digilife.be/quickreferences/QRC/UNIX
%20commands%20reference%20card.pdf
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chmod and tar chmod
There is a nice tutorial here http://www.perlfect.com/articles/chmod.shtml
tar There is a nice tutorial here
http://www.thegeekstuff.com/2010/04/unix-tar-command-examples/
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Running perl on binf.gmu.edu
% ssh binf.gmu.eduPassword: ******
-- Create binf634 directory (don't type stuff in red)% mkdir binf634% cd binf634% ls
-- Copy a file to current directory-- (the "." means :current directory")% cp ~jsolka/public_html/fall15/binf634/bookcode/examples/example4-1.pl .% ls-- This is a one not a l% ls -l
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Running perl on binf.gmu.edu% cat example4-1.pl #!/usr/bin/perl -w# Example 4-1 Storing DNA in a variable, and printing it out
# First we store the DNA in a variable called $DNA$DNA = 'ACGGGAGGACGGGAAAATTACTACGGCATTAGC';
# Next, we print the DNA onto the screenprint $DNA;
# Finally, we'll specifically tell the program to exit.exit;
-- Changing permissions% chmod 755 example4-1.pl
-- Running a perl script% example4-1.pl
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Editing a Perl Script-- Read the Emacs or vi tutorial.-- Make a copy and edit the copy% cp example4-1.pl first.pl% l% vim first.pl-- 1. Change 'print $DNA;' to 'print $DNA, "\n";'-- 2. Now add a comment:# Author: your name% cat first.pl#!/usr/bin/perl -w# Author: Jeff Solka# Example 4-1 Storing DNA in a variable, and printing it out
# First we store the DNA in a variable called $DNA$DNA = 'ACGGGAGGACGGGAAAATTACTACGGCATTAGC';
# Next, we print the DNA onto the screenprint $DNA, "\n";
# Finally, we'll specifically tell the program to exit.exit;
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For Next Week Read Tisdall chapters 1-5.
Be ready to ask questions Be ready to answer questions
HW 1: Write programs as described in the following exercises from "Beginning Perl for Bioinformatics" by Tisdall:
4.3, 4.4, 4.5, 5.2, 5.4 and 5.6 For each exercise, create a perl script called exX.Y.pl, for
example, ex4.3.pl for the first exercise. Submit these via Blackboard
No class next week because of labor day
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Some of the Details
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Alternative Development Environments
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What is Eclipse? Eclipse is a multi-language software development platform
comprising an IDE and a plug-in system to extend it. It is written primarily in Java and is used to develop applications in this language and, by means of the various plug-ins, in other languages as well—C/C++, Cobol, Python, Perl, PHP and more.
The initial codebase originated from VisualAge.[1] In its default form it is meant for Java developers, consisting of the Java Development Tools (JDT). Users can extend its capabilities by installing plug-ins written for the Eclipse software framework, such as development toolkits for other programming languages, and can write and contribute their own plug-in modules. Language packs provide translations into over a dozen natural languages.[2]
Released under the terms of the Eclipse Public License, Eclipse is free and open source software.
http://en.wikipedia.org/wiki/Eclipse_(software)
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What Operating Systems Does Eclipse Run Under? LINUX
MAC OSX
WINDOWS 7 8 10 ?
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Languages Supported by the Eclipse IDE JAVA
Out of the box PERL
Via EPIC library Note one must also have a PERL compiler
PYTHON Via PyDev library
Note one must also have a PYTHON compiler installed
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Advantages and Disadvantages of the Eclipse Development Environment
Advantages Support for a plethora of languages Industrial strength
Used by many professional software developer Has support for configuration management
Disadvantages Can be slow when developing in languages other than JAVA
(may be mere anecdotal evidence)
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Installing Eclipse Under Windows XP - I First make sure that you have a Java Runtime Environment
installed Microsoft Windows XP [Version 5.1.2600](C) Copyright 1985-2001 Microsoft Corp.
C:\Users\jlsol_000>C:"\Program Files\Java\jdk1.7.0_12\bin"\java –version
If you don’t have a JRE installed go to http://www.oracle.com/technetwork/java/javase/downloads/jre8-
downloads-2133155.html
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Installing Eclipse Under Windows XP - II Obtain the Eclipse zipped file from the Eclipse
downloads link at http://www.eclipse.org/downloads/ I believe that I chose this one
Eclipse IDE for Java Developers (163MB) I think that the standard version works also I think that the current version of Eclipse is Mars
Unzip it into an eclipse folder under your windows Program Files directory
In my case here C:\Program Files\eclipse
Note that Eclipse does not modify your system’s registry
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Installing Eclipse Under Windows XP - III Once installed (unzipped)
Double click on the eclipse.exe icon There is a “hello world” java tutorial
There are a number of other tutorials Eclipse3-1.pdf
(http://www.cs.umanitoba.ca/~eclipse/Eclipse3-1.pdf) http://wiki.eclipse.org/
Eclipse_Articles,_Tutorials,_Demos,_Books,_and_More
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Downloading ActiveStates ActivePerl Go here and click on the Windows download link
http://www.activestate.com/activeperl/ I previously set this up using version 5.10 Current version is 5.20.2
Use this self extracting binary to install the program This takes a long time (30 minutes or more, go enjoy your
favorite beverage)
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Installing the Eclipse EPIC Library This is my synopsis of this EPIC webpage tutorial
http://www.epic-ide.org/download.php This is also a helpful site
http://www.epic-ide.org/faq.php Under Eclipse user the Help->Software Updates Tab Switch to the Available Software tab Choose Add Site and choose
http://e-p-i-c.sf.net/updates Tick the newly created site and click the install button
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Creating Your First PERL Program Under the Eclipse IDE - I Under Eclipse go to Window -> Open Perspective ->
Other Choose PERL
Under Eclipse go to Window -> Preferences Click on the PERL + and enter in the full path to the
ActiveStates PERL executable In my case it is "C:\Perl\bin\perl5.10.0.exe"
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Creating Your First PERL Program Under the Eclipse IDE - II Click on File -> New PERL Project
Call it something like HelloWorld Click on File -> New PERL File
Call it something like HelloWorldPerl Left click on this file symbol and make sure its extension is .pl
(Now it should have a camel symbol) Enter in your code
print "Hello from ActivePerl!\n"; Now you should be able to choose Run from the top menu or
left click on the program symbol and choose Run As Perl Local If all goes well a console window with the output
Hello from ActivePerl!
should show up
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Debugging With Eclipse and PERL The Perl PPM package PadWalker has to be installed
before one can debug your PERL programs under Eclipse
Follow the steps on the next two slides to install PadWalker within ActiveStates PERL
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First Find the Package (PadWalker) Find a package. To find a package in the repository: Click the All packages button, Enter text from the package's name or abstract in the
Filter field As text is entered in the Filter field, the list of packages
is automatically updated as the substring match becomes more precise. Click the magnifying glass icon to filter on different meta-data (e.g. Author).
Alternatively, just start typing the name of the package. The Package List will highlight the first package that matches the string you have typed.
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Next Install the Package (PadWalker) Install a package. To install a package from the repository: Click on the desired package in the Package List to
select it. Mark the package by:
Clicking the Mark for install button or, Hitting the "+" key or, Selecting Install <package-name> from the Action menu
or, Right-clicking the selection and choosing Install <package-
name> from the context menu. Click the Run marked actions button or select Run
Marked Actions (Ctrl-Enter) from the File menu. In my case I installed PadWalker 1.7
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Installing PadWalker Via ppm There are other interesting discussions here but they
seem to have been somewhat relegated by the gui-based ActiveStates PERL ppm interface
http://trouchelle.com/perl/ppmrepview.pl
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Editors
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http://www.viemu.com/vi-vim-cheat-sheet.gif
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https://www.gnu.org/software/emacs/refcards/pdf/refcard.pdf
winscp A graphical user interface to scp
Available here
https://winscp.net/eng/download.php
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Notepad ++ A nice editor for windows
Can be configured to execute Perl via the Nppexec plugin
Available here
https://notepad-plus-plus.org
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Suggested Paths Do editing on own system using notepad ++ copy to
binf using scp and run there This is best
Can do development on your own system using Perl and notepad ++
Always test programs on binf
If you are adventurous use Eclipse but it can be confusing to the computer science novice
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