There are many different representations (or, in other words, formats) of DNA and protein sequences (joined with ancillary annotations). Almost every program introduces its "native" format optimized for a particular program needs, and some programs include converters from one format to another. However, some programs read sequences only in one or the other formats. To recognize formats and to be able to interconvert files in different formats is very important for successful data analysis.
There are sets of symbols used to abbreviate nucleotides and amino acids. The characters can be either upper or lower case. The characters enable input of nucleic acid sequences taking full account of any ambiguities in the sequence.
Symbol Meaning
------ -------
A Adenine
G Guanine
C Cytosine
T Thymine
U Uracil
Y pYrimidine (C or T)
R puRine (A or G)
W "Weak" (A or T)
S "Strong" (C or G)
K "Keto" (T or G)
M "aMino" (C or A)
B not A (C or G or T)
D not C (A or G or T)
H not G (A or C or T)
V not T (A or C or G)
X,N,? unknown (A or C or G or T)
O deletion
- deletion
The protein sequences are given by the one-letter code used by the late Margaret Dayhoff's group in the Atlas of Protein Sequences. They are as follows:
Symbol Stands for
------ ----------
A ala
B asx
C cys
D asp
E glu
F phe
G gly
H his
I ileu
J (not used)
K lys
L leu
M met
N asn
O (not used)
P pro
Q gln
R arg
S ser
T thr
U (not used)
V val
W trp
X unknown amino acid
Y tyr
Z glx
* nonsense (stop)
? unknown amino acid or deletion
- deletion
where "nonsense", and "unknown" mean respectively a nonsense (chain termination) codon and an amino acid whose identity has not been determined. The state "asx" means "either asn or asp", and the state "glx" means "either gln or glu" and the state "deletion" means that alignment studies indicate a deletion has happened in the ancestry of this position, so that it is no longer present.
Here are the same one-letter codes tabulated the other way 'round:
Amino acid One-letter code
---------- ---------------
ala A
arg R
asn N
asp D
asx B
cys C
gln Q
glu E
gly G
glx Z
his H
ileu I
leu L
lys K
met M
phe F
pro P
ser S
thr T
trp W
tyr Y
val V
deletion -
nonsense (stop) *
unknown amino acid X
unknown (incl. Deletion) ?
The simplest format for DNA/protein sequences is a FASTA format (or Pearson format). It is used in a variety of molecular biology software. Every sequence in the file starts with "greater than" character (>). That character is followed by an identifier of a sequence (e.g. name, description, gi number) and a carriage return (it is also called a paragraph sign). This line is called a definition line. Everything after carriage return is considered as a sequence. The example of a sequence in FASTA format is shown below:
>gi|2978501|gb|AAC06133.1| vacuolar ATPase proteolipid subunit [Giardia intestinalis] MSSIDSPVAVEKCPAGASFWSMLGQVVAVVFSSIGAAYGTAKAGSGLGV AGLINPAPVTKLTLPVIMAGILSIYGLITSLLINSRVRSYTNGMPLYVS YAHFGAGLCCGLAALAAGLAIGVSGSAAVKAVAKQPSLFVVMLIVLIFS EALALYGLIIALILSTKSADSNFCVNNVNQ
Practical Tip1: The first line might be long and be wrapped around in a text editor or web browser. Often web browsers introduces carriare returns after every line of a text. If you cut and paste the sequence from a web browser, check its contents in a text editor (such as MS Word), and remove any carriage returns. Otherwise, part of the definition line will be interpreted as a sequence.
Practical Tip2: It is very often convenient to put species name in the beginning of the definition line because the programs often take first several symbols of the definition line as the identifiers for the sequences (see examples below).
GenBank format (also called GenBankFlatFile format) is one of the formats that shows the complete information on a sequence entry. Every record in GenBank format consists of 3 parts: the header, the features that describe the annotations on the record, and the sequence sequence itself. Very detailed decription of every field in a GenBank record is available here.
The example of GenBank record is shown below:
LOCUS AF123456 1512 bp mRNA VRT 23-MAR-1999
DEFINITION Gallus gallus testis-specific mRNA sequence.
ACCESSION AF123456
VERSION AF123456.1 GI:4454562
KEYWORDS .
SOURCE chicken.
ORGANISM Gallus gallus
Eukaryota; Metazoa; Chordata; Craniata; Vertebrata; Archosauria;
Aves; Neognathae; Galliformes; Phasianidae; Phasianinae; Gallus.
REFERENCE 1 (bases 1 to 1512)
AUTHORS Nanda,I., Shan,Z.H., Schartl,M., Burt,D.V., Koehler,M.,
Nothwang,H.-G., Gruetzner,F., Paton,I.R., Windsor,D., Dunn,I.,
Engel,W., Staeheli,P., Mizuno,S., Haaf,T. and Schmid,M.
TITLE 300 million years of conserved synteny between chicken Z and human
chromosome 9
JOURNAL Nat. Genet. 21 (3), 258-259 (1999)
MEDLINE 99178258
REFERENCE 2 (bases 1 to 1512)
AUTHORS Shan,Z.H. and Haaf,T.
TITLE Isolation of the Z-linked copy of human DMT1 in chicken and
analyzing its role in developing gonads, further evidence for
evolutionary conservation of sex determining genes
JOURNAL Unpublished
REFERENCE 3 (bases 1 to 1512)
AUTHORS Haaf,T. and Shan,Z.H.
TITLE Direct Submission
JOURNAL Submitted (25-JAN-1999) Max-Planck Institute for Molecular
Genetics, Ihnestr. 73, Berlin 14195, Germany
COMMENT [WARNING] On Dec 23, 1999 this sequence was replaced by a newer
version gi:6633795.
FEATURES Location/Qualifiers
source 1..1512
/organism="Gallus gallus"
/db_xref="taxon:9031"
/chromosome="Z"
/map="Zp21"
/note="testis-specific mRNA"
BASE COUNT 320 a 434 c 473 g 285 t
ORIGIN
1 cccggcgcgg gcaagaagct gccgcgtctg cccaagtgtg cccgctgccg caaccacggc
61 tactcctcgc cgctgaaggg gcacaagcgg ttctgcatgt ggcgggactg ccagtgcaag
121 aagtgcagcc tgatccgccg agcggcaggg gtgatggccg tgcaggttgc actgaggagg
181 cagcaagccc aggaagagga gctggggatc agcgcaccct gtacccctgc ccagtgcccc
241 tgagccagtt gtcaagaaga gcagcagcag cagctcctgt ctcctgcagg acagcagcag
301 cccctgctca ctccacgagc acggtggcag cagcagcagc gagcgcacca ccagagggac
361 ggatgctcat tcaggacatc ccttccatcc ccagcagagg gcacttggag agcacgtctg
421 atttggttgt ggactccacc tactacagca gtttttacca gcatccctgt atccttacta
481 taacaacctg tacaactact cccagtacca aatggcagtg gccactgagt cttcctcaag
541 tgagacaggg ggtacgtttg tagggtcagc catgaaaaac agccttcgaa gcctcccagc
601 aacatacatg tcaagccagt caggaaaaca gtggcagatg aaagggatgg agaaccgcca
661 tgccatgagc tcccagtacc ggatgtgctc ctactacccg cccacctcat acctgggcca
721 gggggttggc agtcccacct gcgtcacaca gatactggcc tcggaggaca ccccctccta
781 ctcagagtcg aaagcgagag tgttttcgcc gcccagcagc caggactcgg gcctggggtg
841 cctgtcgagc agcgagagca ccaagggaga cctggagtgc gagccccacc aagagcccgg
901 cgccttcgcg gtgagcccgg ttcttgaggg cgagtaggcg cggcgtcggg cggctgctgc
961 gcggcgttca ctgttgcctt gttctgttgg gggttgcggg ggggcgttgg gtttcttctt
1021 tccggggcgg ggggggcacg gcggggccgc ggccgggccg gcggggcggg gcggggcggg
1081 acggggcggg gcggagccgc gcgggggccg cagtccgggc cggggccgcc gtcgggtctc
1141 ggcccgctcc cgtcggggcg gagcgtccga cgatcggcct ccacgaaacg cggtgccgtg
1201 atgtgtttgt agtggttcct cgtaggctcc agacgttttc tcctcgtatc gccaaattaa
1261 cgcgttttgc atattacagt tgagtgcctc gacttagatt gcaatataag cggccagcaa
1321 acaagtctca aaaaaaagtt acgtgcgttt ctgcgagtgt tattttgtta agaacggctc
1381 acagtgtcct cttcctgtgt tacagaagcc aacctgaaat gaaactagtc tggaaaaatt
1441 cattgttctc tgtagttgca gctgtacctg aaataaaaat gttattgatg actgaaaaaa
1501 aaaaaaaaaa aa
//
The following formats are the formats of aligned sequences.
One format which is commonly used is PHYLIP format (which comes from PHYLIP package):
6 13 Archaeopt CGATGCTTAC CGC HesperorniCGTTACTCGT TGT BaluchitheTAATGTTAAT TGT B. virginiTAATGTTCGT TGT BrontosaurCAAAACCCAT CAT B.subtilisGGCAGCCAAT CAC
The first line of the input file contains the number of species and the number of characters, in free format, separated by blanks (not by commas). The information for each species follows, starting with a ten-character species name (which can include punctuation marks and blanks), and continuing with the characters for that species.
The sequences can continue over multiple lines; as a consequence, there are two flavors of the format: interleaved and sequential. In sequential format all of one sequence is given, possibly on multiple lines, before the next starts. In interleaved format the first part of the file should contain the first part of each of the sequences, then possibly a line containing nothing but a carriage-return character, then the second part of each sequence, and so on. Only the first parts of the sequences should be preceded by names. Here is a hypothetical example of interleaved format:
5 42 Turkey AAGCTNGGGC ATTTCAGGGT Salmo gairAAGCCTTGGC AGTGCAGGGT H. SapiensACCGGTTGGC CGTTCAGGGT Chimp AAACCCTTGC CGTTACGCTT Gorilla AAACCCTTGC CGGTACGCTT GAGCCCGGGC AATACAGGGT AT GAGCCGTGGC CGGGCACGGT AT ACAGGTTGGC CGTTCAGGGT AA AAACCGAGGC CGGGACACTC AT AAACCATTGC CGGTACGCTT AA
while in sequential format the same sequences would be:
5 42 Turkey AAGCTNGGGC ATTTCAGGGT GAGCCCGGGC AATACAGGGT AT Salmo gairAAGCCTTGGC AGTGCAGGGT GAGCCGTGGC CGGGCACGGT AT H. SapiensACCGGTTGGC CGTTCAGGGT ACAGGTTGGC CGTTCAGGGT AA Chimp AAACCCTTGC CGTTACGCTT AAACCGAGGC CGGGACACTC AT Gorilla AAACCCTTGC CGGTACGCTT AAACCATTGC CGGTACGCTT AA
CLUSTAL (.aln) format was originated in the alignment program CLUSTAL. The file starts with word CLUSTAL. The alignment is written in blocks of 60 residues. Every block starts with sequence names. The example of the alignemnt in CLUSTAL format is shown below:
CLUSTAL X (1.8) multiple sequence alignment
R.sodomens ---------------------------------CAACCUGA-GAGUU-U-GA-U-CCU-G
R.rubrum4 --------------------------------UUCCCUGAA-GAGUU-U-GA-U---U-G
Ag.tumefac -------------------------------CUCAACUUGA-GAGUU-U-GA-U-CCU-G
Ag.rhizog2 --------------------------------UUCCCUGAA-GAGUU-U-GA-U-CCU-G
Rhb.legum4 --------------------------------UUCCCUGAA-GAGUU-U-GA-U-CCU-G
Rhb.legum6 --------------------------------UUCCCUGAA-GAGUU-U-GA-U-CCU-G
Bdr.japon8 --------------------------------UUCCCUGAA-GAGUU-U-GA-U---U-G
* * ***** * ** * * *
R.sodomens GCUC-A-G-AAC-GAAC-GC--U-GGC-GGC-A-GG-C-CU--AACACA-UGCAA---G-
R.rubrum4 GCUC-A-G-GAC-GAAC-GC--U-GGC-GGC-A-GG-C-CU--AACACA-UGCAA---G-
Ag.tumefac GCUC-A-G-AAC-GAAC-GC--U-GGC-GGC-A-GG-C-UU--AACACA-UGCAA---G-
Ag.rhizog2 GCUC-A-G-AAC-GAAC-GC--U-GGC-GGC-A-GG-C-UU--AACACA-UGCAA---G-
Rhb.legum4 GCUC-A-G-AAC-GAAC-GC--U-GGC-GGC-A-GG-C-UU--AACACA-UGCAA---G-
Rhb.legum6 GCUC-A-G-AAC-GAAC-GC--U-GGC-GGC-A-GG-C-UU--AACACA-UGCAA---G-
Bdr.japon8 GCUC-A-G-AGC-GAAC-GC--U-GGC-GGC-A-GG-C-UU--AACACA-UGCAA---G-
**** * * * **** ** * *** *** * ** * * ****** ***** *
There are standalone programs available which allow interconversion b/w different formats. One of such programs is ReadSeq and it is available as online version here.
References:
The Nexus format will be discussed later in class.