Exons:
Relevant bits of information for using the MUSC Shared Resource,
 the BioMolecular Computing Resource (BCR).

Exon 11-The rapid searches.

FASTA answers the question  'are there any sequences in the 
database which are similar to my query sequence?'.  FASTA 
searches do not provide any formal statistical estimate of the 
similarities.  Using the GCG command SHUFFLE which 
substitutes a random sequence with the same composition as 
your query sequence is A way to examine the validity of your 
FASTA searches. FASTA searches allow or gaps. FASTA searches are 
reportedly MORE SENSITIVE than comparable BLAST searches for 
nucleic acid sequences. FASTA will compare a protein sequence 
against a protein database OR compare a nucleotide sequence against 
a nucleotide database. You have the option of running these searches 
interactively (boring) or in batch ( the option or batch is "-batch"). 
You may:

1. Select all or a range of your sequence as the query.
2. Select all of the GenBank database or any subset. (eg GB-PR:*)
3. Select a locally produced database.

ALL databases MUST be identified  properly! The complete form is:

  database: *     >database name , a colon and  sequence name-the 
asterisk is a "wild card".

examples
gb_pr:*              >searches ALL primate sequences in Genbank
gb_pr:hum*      >searches ALL primate sequences in Genbank 
which begin with the letters "hum"
ides:*                 >searches ALL of the LOCAL database  called ide
(see the DATASET command and Exon _12)                    

TFASTA answers the question 'what implied protein sequences 
in a nucleotidedatabase match my protein query? 'TFASTA 
REQUIRES a protein query sequence. ALL six reading frames of 
the nucleotide database are searched. TFASTA has a LARGE 
overhead and should probably NOT be run interactively!


The BLAST searches: Blast searches are presently about the 
fastest means to answer the question "what sequences are similar 
to my query sequence?"

From GCG
Type "blast" at the prompt. Enter a query sequence and select a 
database to search. From these bits of information the GCGBlast
interface determines which program to run, bundles your query 
sequence and sends a message to the NCBI BLAST server in 
Bethesda, MD.

program        query type        database type
BLASTP         protein           protein
TBLASTN        protein           nucleic acid
BLASTX         nucleic acid      protein
BLASTN         nucleic acid      nucleic acid


The GCG v8.0 allows BLAST searches of local databases.
These databases are created with the program "TOBLAST". 
Notifying the system of the existence of your local database 
requires that you complete several tasks.

1. Create the database with the "toblast" program. TOBLAST can 
create a 'blastable" database (in a binary form) from any list of 
sequences. Output from WORDSEARCH and STRINGSEARCH
 are particularly useful input files for TOBLAST.

2. Fetch ( a GCG command) the file called "blast.sdbs". Type 
"fetch blast.sdbs".
The system then writes the ascii file blast.sdbs to your current 
directory (it is advisable to write the file to your personal root 
directory).

3. Edit the blast.sdbs file to point to your newly minted local 
blastable database(s) by adding a line which  includes a path,
 a name, a type (eg protein or nucleic acid) , and a comment. 
Example lines:

/home/hazards/project1/ides n A nucleotide database
/home/hazards/project2/ICE p A protein database

You must add a name for each local database you create!

4. At the time you evoke the GCG BLAST interface you must add 
the option
"-dat3=/path/blast.sdbs"
This tells the system where to find the file that points it to ALL of 
your local databases.

After indicating your  query sequence you will see a screen of 
databases preceded by numbers. At the bottom of the screen it will 
say "local databases" and below that will be numbers of  each of 
your databases. Selecting the number of one of your local 
databases will direct the GCG interface to use your query sequence 
and search the indicated local database.

The UNIX BLAST programs perform all the same functions as the 
GCG interface EXCEPT that they may NOT search a local database. 
But you may still invoke the BLAST3 search from the UNIX 
command line. BLAST3 may NOT be selected from the GCG 
interface.

These programs require that any query sequences be in the 
so-called fasta format. Any GCG file may be converted to  fasta 
format with the command "tofasta". Any sequence retrieved from 
Network Entrez may be saved (an Entrez option) in fasta format 
directly.

The UNIX BLAST command line is as follows:

program   database     query sequence(fasta format)  options  ">" 
character   outfilename and "&" character.

eg

blastx   nr      ides.tfa   -x    >    ides.blastx   &

This line says that the program blastx is being called, the 
non-redundant PROTEIN database is selected ( selection of 
database is a consequence of selecting the program; there is also 
a non-redundant NUCLEIC ACID database), ides.tfa is a nucleic 
acid sequence in "fasta" format, -x 
selects a repeat filter, the ">" character is a UNIXism pointing to an 
output file named ides.blastx and the "&" character is another 
UNIXism which runs the program in background and frees you to 
continue issuing commands in the current login shell while the 
search is in progress.

MAC BLAST
The HYPERBLAST and BLAST1.4 programs are BLAST packages 
which accept sequences, configure a BLAST query, send the query 
and retrieve the results to your MAC. The programs are available 
(there may also be other sites) via  FTP at:

amber.mgh.harvard.edu
Use "blast" as the username  and "internet" as the password. 

There is a DOS BLAST program which accepts a query sequence, 
allows selection of the database, bundles the information, sends it to 
Bethesda, and retrieves the results.

There is a WINDOWS program which CONFIGURES a query only. 
Sending the file is your responsibility.