Welcome to lecture 2:
Feeling at home in *nix
IGERT – Sponsored Bioinformatics Workshop Series
Michael Janis and Max Kopelevich, Ph.D.
Dept. of Chemistry & Biochemistry, UCLA
Last time…
• We covered a bit of material…
• Try to keep up with the reading – it’s all in there!
• How’s it coming along?
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BioKnoppix
Remote logins, navigation
Unix / linux concepts?
General questions?
The CLI and YOU
Most of bioinformatics is accomplished through command-line tools
• Command line interaction is easily batched
• Command line interaction is easily integrated
• Command line interaction is a form of PROGRAMMING
• It’s therefore worthwhile to become familiar with your *nix environment in a
non-graphical interface
Commands
• In Bioinformatics, we are mostly concerned with TEXT
PROCESSING – the CLI is well suited for this type of work
• Specific commands are used to perform functions in the
shell
• Each command is itself a program and takes command
line arguments
– The syntax order is program [-options] filename
• For help on a specific command type:
man command; apropos topic; command --help
Some review of system tools
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Who
W
Uname
Pwd
Find
Top
Another example of a pipe
file
Command 1
(cut)
Pipe
cut –d: -f1 < /etc/passwd
|
Command 2
(sort)
Stdout
sort
• The file /etc/passwd stores information about
user’s accounts on the system
• Let’s get a sorted listing of all user names
Example: redirecting STDOUT
STDIN
Command
Or
Program
STDOUT
OUTPUT_FILE
cut –d: -f1 < /etc/passwd
|
sort
> output_file
more output_file
“redirection operator”
Process Control
• Each specific job / command is called a process
• Each process runs in a shell
– BEFORE: prompt available
– DURING: prompt NOT available
– AFTER: prompt available
• Control keys
– CTRL-C -> stop current command
– CTRL-D -> end of input
Two Ways to monitor Processes
• “top”
– Lists all jobs
– Uses a table format
– Dynamically changes
• “ps”
– man ps
– static content
– Command options
What are you doing, Dave?
Background / Foreground
• Commands running in foreground prevent
prompt from being used until command
completes
• Commands can also run in BACKGROUND
• “Backgrounded” commands DO NOT AFFECT
the prompt
Two Ways to Background jobs
• “&”
– Running a command with
“&” automacically sends it
to the background
– Backgrounded commands
return the prompt
• “bg”
– Once a command is run
from the prompt
– Stop the command
– Then background it
• Starts the command again
• Returns the prompt for
use
File System Navigation
• Absolute filepaths begin with the root ‘/’
• Relative filepaths don’t have a preceding slash; they begin from the
cwd
• What is the absolute path to cd from john to mary?
• What is the relative path to cd from john to mary?
• Once you are in mary, and your username is john, what are two ways
to return to your home directory?
The society for anti-defamation of
computer mouses opposes this slide
• There’s very little reason to leave the CLI
• Most tasks can be written within the shell
• The user-friendliness becomes self-limiting
Let’s take an example…
• Suppose you wanted to do some biological
analysis – like motif searching through a
database of biological sequences… What do you
need to do this?
– You need to retrieve the sequences
– You need to describe the motif
– You need to search the sequences
I want to search for zinc-finger
motifs genomically in yeast (S.c.)
• I’m going to need the genomic sequence for
Saccharomyces cerevisiae
(http://www.yeastgenome.org)
• I’m going to need the motif that describes the
zinc finger I’d like to search for (ProSite).
• I’m going to need do do this search many times
across every chromosome.
A brief overview of some databases /
biological information repositories
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NCBI
Genome-specific databases (SGD…)
SMD http://genome-www5.stanford.edu/
The Stanford Microarray Database. Repository of microarray analysis from a wide variety.
PROSITE http://au.expasy.org/prosite/
Used to rapidly search your protein sequences for catalogued motifs.
SWISSPROT http://www.ebi.ac.uk/swissprot/
SWISSPROT is a "one stop shop" for protein sequence information. Use it to extend your
knowledge of your proteins.
PDB: The Protein Databank http://www.rcsb.org/pdb/
The Protein Data Bank is the single worldwide archive of structural data of biological
macromolecules. Structure implies function in general.
PFAM: http://www.sanger.ac.uk/Software/Pfam/search.shtml
This database is a collection of protein motifs.
PRODOM http://protein.toulouse.inra.fr/prodom/current/html/home.php
PRODOM is similar to PFAM in that it is a set of curated protein domain families. However, the
underlying computational engine is different.
BLOCKS http://blocks.fhcrc.org/
Blocks are multiply aligned ungapped segments corresponding to the most highly conserved
regions of proteins. The blocks for the Blocks Database are made automatically by looking for the
most highly conserved regions in groups of proteins documented in InterPro.
COG http://www.ncbi.nlm.nih.gov/COG/
COG stands for Clusters of Orthologous Groups of proteins. This is a tool for phylogenetic
classification of proteins encoded in complete genomes. COGs were delineated by comparing
protein sequences encoded in complete genomes, representing major phylogenetic lineages.
Retrieving data
Retrieving data
• You don’t have to leave the CLI. Really.
– If you need to do something, chances are there’s a
utility to do so
– Debian is your friend (search packages FIRST!!!)
Introducing wget:
>wget ftp://genomeftp.stanford.edu/pub/yeast/data_download/protein_info/hypothetical
_peptides/*.gz
Of course you can use ftp:
>ftp genome-ftp.stanford.edu
-login anonymous; use your email address as passwd
-traverse filesystem like any linux CLI
-bin, get, prompt, mget…
A note about file archives
• Most files will be compressed. Usually using
gunzip.
• Most files will be agglomerative, using TAR.
Introducing gunzip:
>gunzip *.gz
Introducing tar (tape archive):
>tar –xvf *.tar
Or to create a tar
>tar –cvf output.tar *.*
A brief note about the biological
file format called FASTA
• In bioinformatics, FASTA format is a file format used to exchange
information between genetic sequence databases. Its format looks like
this:
• >SEQUENCE_1 ;comment line 1 (optional)
MTEITAAMVKELRESTGAGMMDCKNALSETNGDFDKAVQLL
REKGLGKAAKKADRLAAEGLVSVKVSDDFTIAAMRPSYLSYE
DLDMTFVENEYKALVAELEKENEERRRLKDPNKPEHKIPQFAS
RKQLSDAILKEAEE
• It consists of a header line (beginning with a '>') which gives a name
and/or a unique identifier for the sequence. Many different sequence
databases use FASTA files.
• After the header line and comments, one or more sequence lines may
follow. Sequences may be protein sequences or DNA sequences
– they must be shorther than 80 characters and can contain gaps or
alignment characters
• FASTA format files often have file extensions like .fa or .fsa
• The simple format of FASTA files makes them easy to manipulate
using text processing tools and scripting languages like Perl.
*From http://en.wikipedia.org/wiki/Fasta_format
ProSite motif
Describing the motif - GREP
• “GREP” searches contents of a file or directory
of files
– “Get Regex” – uses regular expressions
– File wildcards can be used like with ls
• grep 1sq ~/DATA/*.CEL -> array type used
– We explored this last time (briefly!)
Regular expressions
•
A regular expression, often called a pattern, is an
expression that describes a set of strings. They are
usually used to give a concise description of a set,
without having to list all elements.
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–
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For example, the set containing the three strings Mike, Mark, and
Matt can be described by the pattern “M((ike|(ark|att))?)"
Alternatively, it is said that the pattern “M((ike|(ark|att))?)"
matches each of the three strings.
There are usually multiple different patterns describing any
given set. Most formalisms provide the following operations to
construct regular expressions.
Formalisms of regular expressions
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alternation
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A vertical bar separates alternatives. For example, "gray|grey" matches grey or
gray.
grouping
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Parentheses are used to define the scope and precedence of the operators. For
example, "gray|grey" and "gr(a|e)y" are different patterns, but they both
describe the set containing gray and grey.
quantification
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A quantifier after a character or group specifies how often that preceding
expression is allowed to occur. The most common quantifiers are ?, *, and +:
?
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The question mark indicates that the preceding character may be present at most once.
For example, "colou?r" matches color and colour.
*
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+
•
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The asterisk indicates that the preceding character may be present zero, one, or more
times. For example, "0*42" matches 42, 042, 0042, etc.
The plus sign indicates that the preceding character must be present at least once. For
example, "go+gle" matches the infinite set gogle, google, gooogle, etc. (but not ggle).
These constructions can be combined to form arbitrarily complex
expressions, very much like one can construct arithmetical expressions from
the numbers and the operations +, -, * and /.
*From http://en.wikipedia.org/wiki/Regular_expression
The real world is fuzzy and
complex…
• What if we just want to search for a string in the format
of a phone number;
• E.g.
825 8901
213 487 0353
No area code
Area code
• Obviously we can’t check for each possible phone number
(some 1010 possibilities makes for a very long set of
statements…).
This is where regular expressions
come in…
• Regular expressions describe
generalised patterns of strings instead
of exact strings.
>grep /([0-9]{3} ){0,1}[0-9]{3} [0-9]{4}/) filename
• (clearly this is a little more complex as
an example…)
Special characters
(‘metacharacters’)
‘.’ is a wildcard and matches any character
>grep ‘.ed’ filename
If file contains
-will find
If file contains
-will find
If file contains
-will not find
If file contains
-will find
“bed”
“red”
“head”
“edward”
Special characters
(‘metacharacters’)
‘*’ means ‘zero or more of the previous character’.
>grep ‘be*d’ filename
If file contains
-will find
If file contains
-will not find
If file contains
-will find
If file contains
-will find
“bed”
“red”
“beeeed”
“bd”
Special characters
(‘metacharacters’)
‘+’ means ‘one or more of the previous character’.
>grep ‘be+d’ filename
If file contains
-will find
If file contains
-will not find
If file contains
-will find
If file contains
-will not find
“bed”
“red”
“beeeed”
“bd”
Start and end of line
‘^’ is designates the start of the line, ‘$’ the end.
>grep ‘bed’ filename
If file contains “bed”
-will find
If file contains “bedbed”
-will find
If file contains “xxxbedxxx”
- will find
>grep ‘^bed$’ filename
Iff file contains “bed” on
line by itself
-will find
If file contains “bedbed”
-will not find
If file contains
“xxxbedxxx” – will not find
Grouping with parentheses
Parentheses group characters
>grep ‘(bed)+’ filename
If file contains “bed”
-will find
If file contains “bedbed”
-will find
If file contains “beddd”
-will not find
Character classes
• The square brackets are used to denote whole
groups of characters
>grep ‘[brf]ed’ filename
If file contains “bed”
-will find
If file contains “red”
-will find
If file contains “led”
-will not find
Character classes (cont)
• A hyphen designates a range:
>grep ‘[a-z]ed’ filename
If file contains “bed”
-will find
If file contains “fed”
-will find
If file contains “Bed”
-will NOT find (why not?)
Character class shortcuts
• Some character classes are so common there are
in-built shortcuts:
– [0-9]
– [A-Za-z0-9]
– [\f\t\n\r ]
=
=
=
\d
\w
\s
Quantifying
• Curly brackets quantify repeats better than ‘*’
(0+) or ‘+’ (1+)
a{3,5}
=
three, four or five ‘a’’s.
>grep ‘la{3,5}’
If file contains “laaaad”
-will find
If file contains “laaaaaaad”
-will not find
Referencing
• Back-slashes match the substring previously
matched by the nth parenthesized subexpression
of the regular expression.
– The back-reference is denoted `\n', where n is a single
digit
>grep ‘(a)\1’
If file contains “laaaad”
-will find
If file contains “lad”
-will not find
Back to our ProSite motif…
• We can use regular expressions to describe the
motif
– The motif is actually a REGULAR EXPRESSION!
>grep -n –E -–color –B2
‘C.{2}C.{4,8}[RHDGSCV][YWFMVIL].[CS].{2,5}[CHEQ].[DNSAGE
][YFVLI].[LIVFM]C.{2}C *.fsa
chr04.peptides.20040928.fsa-4202->Annotated|04:1356055:1357359| frame 1; YDR448W/ADA2;
Verified; this gene contains 1 exon
chr04.peptides.20040928.fsa:4203:MSNKFHCDVCSADCTNRVRVSCAICPEYDLCVPCFSQGSY
TGKHRPYHDYRIIETNSYPILCPDWGADEELQLIKGAQTL
Did it work?
Let’s try this…
• Download the genomic DNA sequence from SGD
• Search for any variant of the TATA – box
promoter
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TATAAA
TATAAT
TATATT
TAATAA
TAATAT
More more more
• Many MS tools allow for wildcard searching
• The shell allows variables; interpolation; control
structures
– For example, attempt to find a palindrome of length 4
within genomic sequences (hint: use backreferences!)
– Variables allow for persistence and control structures
>myVar=`grep -n –E -–color
‘C.{2}C.{4,8}[RHDGSCV][YWFMVIL].[CS].{2,5}[CHEQ].[DNSAGE
][YFVLI].[LIVFM]C.{2}C *.fsa`
[email protected]:~$ echo $myVar
chr04.peptides.20040928.fsa:4203:MSNKFHCDVCSADCTNRVRVSCAICPEYDLCVPCFSQGSY
TGKHRPYHDYRIIETNSYPILCPDWGADEELQLIKGAQTL
A better variable interpolation
• The variable is allowed to change
• We can set the variable to the Prosite Pattern
[email protected]:~$
myVar=C\.{2}C\.{4,8}[RHDGSCV][YWFMVIL]\.[CS]\.{2,5}[CHEQ]\.[DNSAGE][YFVLI]\.[LIVF
M]C\.{2}C
[email protected]:~$ echo $myVar
C.{2}C.{4,8}[RHDGSCV][YWFMVIL].[CS].{2,5}[CHEQ].[DNSAGE][YFVLI].[LIVFM]C.{2}C
[email protected]:~$ grep -n -E --color $myVar *.fsa
chr04.peptides.20040928.fsa:4203:MSNKFHCDVCSADCTNRVRVSCAICPEYDLCVPCFSQGSY
TGKHRPYHDYRIIETNSYPILCPDWGADEELQLIKGAQTL
Variables can be overwritten
• The variable is allowed to change
• We can set the variable to the Prosite Pattern
[email protected]:~$ function afun {
> for i in 1 2 3 4 5
> do
> echo $i
> echo $myVar
> done
>}
[email protected]:~$ afun
1
C.{2}C.{4,8}[RHDGSCV][YWFMVIL].[CS].{2,5}[CHEQ].[DNSAGE][YFVLI].[LIVFM]C.{2}C
2
C.{2}C.{4,8}[RHDGSCV][YWFMVIL].[CS].{2,5}[CHEQ].[DNSAGE][YFVLI].[LIVFM]C.{2}C
3
C.{2}C.{4,8}[RHDGSCV][YWFMVIL].[CS].{2,5}[CHEQ].[DNSAGE][YFVLI].[LIVFM]C.{2}C
4
C.{2}C.{4,8}[RHDGSCV][YWFMVIL].[CS].{2,5}[CHEQ].[DNSAGE][YFVLI].[LIVFM]C.{2}C
5
C.{2}C.{4,8}[RHDGSCV][YWFMVIL].[CS].{2,5}[CHEQ].[DNSAGE][YFVLI].[LIVFM]C.{2}C
Functions
• What if we wanted to search every ProSite
pattern against our genomic database?
• We’d have to repeatedly do our search
– This is called a loop
– We have to write this so the computer knows exactly
what to repeat, how many times to repeat, and where
to find the next ProSite pattern to match
– We would store the what and where in VARIABLES
– We would utilize a CONTROL STRUCTURE to
handle the how…
Control structures
• All out programs so far have run from start to
finish. Each line has been executed in turn.
• What if we only want to run some lines some of
the time?
• This is where control structures come in.
Control structures
• Programming languages generally have a
number of control structures.
• Basic structures:
– if
– while
– for & foreach
• There are others (e.g. unless)
‘for’ example
>afunction() {
for i in 1 2 3 4 5
do
echo "Looping ... number $i"
done
}
Variables can interpolated
• The command is substituted from the system
• It’s like a pipe, but we are allowed to operate
[email protected]:~$ afun() {
> myvar=$(ls -1 *.fsa)
> for i in $myvar
> do
> echo $i
> done
>}
[email protected]:~$ afun
chr01.fsa
chr01.peptides.20040928.fsa
chr02.peptides.20040928.fsa
chr03.peptides.20040928.fsa
chr04.peptides.20040928.fsa
chr05.peptides.20040928.fsa
chr06.peptides.20040928.fsa
chr07.peptides.20040928.fsa
chr08.peptides.20040928.fsa
chr09.peptides.20040928.fsa
chr10.peptides.20040928.fsa
chr11.peptides.20040928.fsa
…
The ‘while’ control structure
(combined with opening files)
• The ‘while’ control stucture keeps looping while a
given condition is satisfied
• ‘while’ and open files go together very well:
[email protected]:~$ afun() {
> while read f
> do
> echo $f
> done
>}
[email protected]:~$ afun < chrmt.peptides.20040928.fsa
>Notannotated|mt:385:459| frame 1
MNYILLLLLIKLLIIINMKLIKIL
…
Editors
• Shell programming is like a batch file
– Commands are linked together in a procedure
– The procedure is accessed via a file
• We need an editor that will allow us to construct
that file
–
–
–
–
–
We’ll use Emacs (or you can use vi, pico, …)
Comprehensive, extensible working environment
Complete (arguable!) IDE
Integration
Extensible (elisp)
Emacs
• Invoking Emacs is easy: emacs –nw filename
• In many cases, Emacs will work out the mode
appropriate for your file (.cpp, .pl, etc…)
– The mode allows Emacs to become sensitive to the task
– There is a biomode for reverse complement, etc….
– You can write your own!
• Emacs has many tools
– Search, replace, cut, paste, mail…
– File navigation, ftp, remote shells…
The Emacs survival guide
•
•
•
•
•
•
Notation
– Emacs uses the control key and escape key heavily. We write it like this:
C-x Pronounced "Control-x“
– Hold down the Ctrl key (usually in the lower left corner of the keyboard) while pressing the x
key.
– Both Ctrl and x must be down at the same time. M-x Pronounced "Meta-x"
Press the Esc key (usually in the upper left corner of the keyboard), release it, then press the x
key.
– Esc and x should not be down at the same time. So C-x C-f means hold down the control key,
then type x and then f while holding it down. (This is the command to load a file into emacs).
Typing
– Just type. All the regular keys, arrow keys, delete, backspace, and page up/down keys should
work. Alternatively, you can try these commands: C-f cursor forward, C-b cursor back, C-p
previous line, C-n next line, M-v page up, C-v page down.
Exiting
– Type C-x C-c. If you have any unsaved work, emacs will ask you if you want to save it. Type y.
Other commands
– Most control or escape sequences are commands. Usually a prompt appears in the command
line at the bottom of the window. Here are a few:
– C-x C-f Load file, prompt for filenameC-x C-s Save file without exiting C-x C-c Exit, prompt
to save files C-s Search forward, prompt for search string C-r Search backward, prompt for
search string C-h ?Show help options, prompt for choice C-h t Start emacs tutorial If you
make a mistake or change your mind you can always escape:
C-g
– Abandon command and resume typing
Command line editing
• Learning the keybindings can be difficult
– But it will increase your speed
– Faster than using a mouse
– Transferable! The keybindings for command line
editing from Emacs is the default set of commands for
line editing in the Bash Shell!
Let’s try it…
• Open up the file that we found contained the
ProSite Motif
• Open a second window
• Goto the line that contains the motif (hint: use
grep with –n!)
• Copy and paste that line into a new file
• Save and close that file
AWK is your pre-perl friend
•
•
•
•
Use to print a subset of fields
Default field delimiter is “ “ (white space)
Useful for grabbing a subset of fields
Useful for rearranging fields
field1 filed2 field3 field4 . . .
$1 $2
$3
$4 . . . .
pipe
Using AWK
| awk –F” “ ‘{print $1}’
| awk –F” “ ‘{print $1”
“$2}’
| awk –F” “ ‘{print $1”\t”$2}’
\t = TAB
\n = newline
Overwrite versus Append
• > OVERWRITE – delete and replace
• >> APPEND – add to end of existing file
Example: microarray data
tracking
• grep 1sq ~/DATA/*.CEL (gives array info)
• grep 1sq ~/DATA/*.CEL | awk ‘{print $12}’ gives
array type only
• grep 1sq ~/DATA/*.CEL | awk ‘{print $12}’ >
arrayTypes.txt (store results in file)
• ls ~/DATA/*.DAT | wc (gives a count)
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Discovering conserved DNA