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#!/usr/bin/perl
use strict;
use warnings;
# gbf2tbl.pl
# NCBI is supplying this Perl script for the convenience of GenBank
# submitters who record their genome annotation in a format similar to
# GenBank or EMBL reports. The gbf2tbl.pl parser allows some variation from
# the official strict flat file conventions. Additional idiosyncrasies in
# format can be supported by modification of the script. Any suggestions for
# improvements or corrections are welcome. However, given the number of
# possible variations of local data files and conditions, NCBI cannot
# promise to implement them all and cannot be expected to troubleshoot local
# installations. This script is being supplied as is, but you are always
# welcome to make any changes or improvements yourself.
#
# The script produces annotated FASTA and 5-column feature table files,
# which are then read by tbl2asn to complete the submission preparation
# process. Submitters are expected to provide data that meet the annotation
# criteria for acceptance by GenBank. This includes annotating coding
# regions and structural RNAs, providing a gene feature with a unique
# /locus_tag qualifier for each, and confirming that all CDS features
# translate into proteins without error.
#
# For convenience, tbl2asn also runs the NCBI sequence record validator,
# which checks the resulting record for many kinds of errors and
# inconsistencies. Even syntactically correct records will be rejected if
# they do not contain information required by GenBank policy. Examining the
# validator output from tbl2asn and correcting the underlying data is
# critical to ensure that submissions are acceptable.
#
# Existing examples from several annotation pipeline outputs have been used
# to build the parser and have it accommodate information as presented.
# Additional conversions are described below:
#
# If the ACCESSION is blank or "unknown", the sequence identifier is taken
# from the LOCUS line.
#
# If the source feature is missing, the taxonomic name is taken from the
# ORGANISM line.
#
# Feature keys or qualifiers that are not present in the INSDC feature table
# document are reported in a .err file, and are not added to the 5-column
# feature table. See http://www.insdc.org/documents/feature_table.html for
# details.
#
# A few non-standard qualifiers are allowed and are converted to the
# appropriate field in the output file. These include codon_recognized,
# go_component, and region_name.
#
# Feature intervals that refer to 'far' locations, i.e., those not within
# the cited record and which have an accession and colon, are suppressed.
# Those rare features (e.g., trans-splicing between molecules) must be
# annotated later using Sequin.
#
# The genetic code in the FASTA definition line, necessary for proper
# translation of protein coding regions, is taken from a CDS /transl_table
# qualifier.
#
# A companion script, tblfix.pl, can perform various data conversions on the
# resulting feature table files, with the conversion function specified by a
# command line argument.
# Script to convert pseudo-GenBank or pseudo-EMBL files to FASTA and
# 5-column feature table files suitable for submission to NCBI using
# the tbl2asn or Sequin programs.
my $gbffile = shift or die "Must supply input filename\n";
open (my $GBF_IN, $gbffile) or die "Unable to open $gbffile\n";
my $base = $gbffile;
if ($base =~ /^(.+)\.gbf$/ || $base =~ /^(.+)\.gbk$/ || $base =~ /^(.+)\.gb$/ ||
$base =~ /^(.+)\.embl$/ || $base =~ /^(.+)\.emb$/ || $base =~ /^(.+)\.eb$/ ||
$base =~ /^(.+)\.art$/) {
$base = $1;
}
open (my $FSA_OUT, ">$base.fsa") or die "Unable to open sequence output file\n";
open (my $TBL_OUT, ">$base.tbl") or die "Unable to open feature table output file\n";
open (my $ERR_OUT, ">$base.err") or die "Unable to open error output file\n";
# define global variables
my $has_errors = 0;
my $line_number = 0;
# state variables for tracking current position in flatfile
my $in_seq;
my $in_feat;
my $in_key;
my $in_qual;
my $current_key;
my $current_loc;
my $current_qual;
my $current_val;
my $organism;
my $topology;
my $is_source;
my $is_translation;
my $transl_table;
my $thisline;
my $curr_seq;
my $locus;
my $accn;
my $is_order;
my $organism_ok;
my $printed_heading;
# subroutine to clear state variables for each flatfile
# start in in_feat state to gracefully handle missing FEATURES/FH line
sub clearflags {
$in_seq = 0;
$in_feat = 1;
$in_key = 0;
$in_qual = 0;
$current_key = "";
$current_loc = "";
$current_qual = "";
$current_val = "";
$organism = "";
$topology = "";
$is_source = 0;
$is_translation = 0;
$transl_table = 1;
$thisline = "";
$curr_seq = "";
$locus = "";
$accn = "";
$is_order = 0;
$organism_ok = 0;
$printed_heading = 0;
}
# hashes for confirming legal feature keys and legal qualifier names
my %legal_keys = ();
my %legal_quals = ();
sub createlists {
my @keys = qw/3clip 3UTR 5clip 5UTR 10_signal35_signal
allele attenuator C_region CAAT_signal CDS centromere
conflict D_loop D_segment enhancer exon gap GC_signal
gene iDNA intron J_segment LTR mat_peptide misc_binding
misc_difference misc_feature misc_recomb misc_RNA
misc_signal misc_structure mobile_element modified_base
mRNA mutation N_region ncRNA old_sequence operon oriT
polyA_signal polyA_site precursor_RNA preprotein preRNA
prim_transcript primer_bind promoter protein_bind RBS
rep_origin repeat_region repeat_unit rRNA S_region
satellite scRNA sig_peptide site_ref snoRNA snRNA
source stem_loop STS TATA_signal telomere terminator
tmRNA transit_peptide tRNA unsure V_region V_segment
variation virion/;
foreach my $thiskey (@keys) {
$legal_keys{lc($thiskey)} = 1;
}
my @quals = qw/allele anticodon artificial_location
bio_material bound_moiety cell_line cell_type
chloroplast chromoplast chromosome citation clone_lib
clone codon_start codon collected_by collection_date
compare cons_splice country cultivar culture_collection
cyanelle db_xref dev_stage direction EC_number ecotype
environmental_sample estimated_length evidence
exception experiment focus frequency function gap_type
gdb_xref gene_synonym gene germline haplogroup haplotype
identified_by inference insertion_seq isolate
isolation_source kinetoplast lab_host label lat_lon
linkage_evidence locus_tag macronuclear map mating_type
metagenomic mitochondrion mobile_element_type
mobile_element mod_base mol_type ncRNA_class note
number old_locus_tag operon organelle organism partial
PCR_conditions PCR_primers phenotype plasmid
pop_variant product protein_id proviral pseudo
pseudogene rearranged replace ribosomal_slippage
rpt_family rpt_type rpt_unit_range rpt_unit_seq
rpt_unit satellite segment sequenced_mol serotype
serovar sex specific_host specimen_voucher
standard_name strain sub_clone sub_species sub_strain
tag_peptide tissue_lib tissue_type trans_splicing
transcript_id transgenic transl_except transl_table
translation transposon UniProtKB_evidence usedin
variety virion go_component go_function go_process
codon_recognized bond_type gene_desc gene_syn prot_desc
prot_note region_name site_type/;
foreach my $thisqual (@quals) {
$legal_quals{lc($thisqual)} = 1;
}
}
# recursive subroutine for parsing flatfile representation of feature location
sub parseloc {
my $subloc = shift (@_);
my @working = ();
if ($subloc =~ /^join\((.+)\)$/) {
my $temploc = $1;
my @items = split (',', $temploc);
foreach my $thisloc (@items) {
if ($thisloc !~ /^.*:.*$/) {
push (@working, parseloc ($thisloc));
}
}
} elsif ($subloc =~ /^order\((.+)\)$/) {
$is_order = 1;
my $temploc = $1;
my @items = split (',', $temploc);
foreach my $thisloc (@items) {
if ($thisloc !~ /^.*:.*$/) {
push (@working, parseloc ($thisloc));
}
}
} elsif ($subloc =~ /^complement\((.+)\)$/) {
my $comploc = $1;
my @items = parseloc ($comploc);
my @rev = reverse (@items);
foreach my $thisloc (@rev) {
if ($thisloc =~ /^([^.]+)\.\.([^.]+)$/) {
$thisloc = "$2..$1";
}
if ($thisloc !~ /^.*:.*$/) {
push (@working, parseloc ($thisloc));
}
}
} elsif ($subloc !~ /^.*:.*$/) {
push (@working, $subloc);
}
return @working;
}
#subroutine to print next feature key / location / qualifier line
sub flushline {
if ($printed_heading == 0) {
if ($accn eq "" || $accn =~ /^unknown.*/) {
$accn = $locus;
}
# report identifier
print $FSA_OUT ">$accn";
print $TBL_OUT ">Feature $accn\n";
$printed_heading = 1;
}
if ($in_key == 1) {
if (! $legal_keys{lc($current_key)}) {
print $ERR_OUT "Bad feature\tLine $line_number\t$current_key\n";
$has_errors = 1;
} elsif ($is_source == 0) {
# parse join() order() complement() ###..### location
$is_order = 0;
my @theloc = parseloc ($current_loc);
# convert number (dot) (dot) number to number (tab) number
foreach my $thisloc (@theloc) {
if ($thisloc =~ /^([^.]+)\.\.([^.]+)$/) {
$thisloc = "$1\t$2";
} elsif ($thisloc =~ /^(.+)\^(.+)$/) {
$thisloc = "$1\^\t$2";
} elsif ($thisloc =~ /^([^.]+)$/) {
$thisloc = "$1\t$1";
}
}
#print feature key and intervals
my $first = shift (@theloc);
print $TBL_OUT "$first\t$current_key\n";
foreach my $thisloc (@theloc) {
print $TBL_OUT "$thisloc\n";
}
if ($is_order == 1) {
# generate order qualifier to force use of order instead of join
print $TBL_OUT "\t\t\torder\n";
}
}
} elsif ($in_qual == 1) {
if (! $legal_quals{lc($current_qual)}) {
print $ERR_OUT "Bad qualifier\tLine $line_number\t$current_qual\n";
$has_errors = 1;
} elsif (! $legal_keys{lc($current_key)}) {
$has_errors = 1;
} elsif ($is_source == 1) {
if ($current_val eq "") {
print $FSA_OUT " [$current_qual=]";
} else {
print $FSA_OUT " [$current_qual=$current_val]";
if ($current_qual eq "organism") {
$organism = "";
$organism_ok = 1;
}
}
} elsif ($current_qual ne "translation") {
if ($current_val eq "") {
print $TBL_OUT "\t\t\t$current_qual\n";
} else {
print $TBL_OUT "\t\t\t$current_qual\t$current_val\n";
}
}
}
}
# initialize flags and lists at start of program
clearflags ();
createlists ();
# main loop reads one line at a time
while ($thisline = <$GBF_IN>) {
$thisline =~ s/\r//;
$thisline =~ s/\n//;
$line_number++;
if ($thisline =~ /^LOCUS\s+(\S*).*$/ ||
$thisline =~ /^ID\s+(\S*);.*$/ || $thisline =~ /^ID\s+(\S*)\s+SV\s+\d+;.*$/) {
# record locus
$locus = $1;
if ($thisline =~ /^.*(linear).*$/ || $thisline =~ /^.*(circular).*$/) {
$topology = $1;
}
} elsif ($thisline =~ /^ACCESSION\s*(\S*).*$/ || $thisline =~ /^AC\s*(.*);.*$/) {
# record accession
$accn = $1;
} elsif ($thisline =~ /^ {1,3}ORGANISM\s+(.*)$/ || $thisline =~ /^OS\s+(.*)$/) {
# record organism
$organism = $1;
if ($organism =~ /^([^(]*) \(.*\)/) {
$organism = $1;
}
} elsif ($thisline =~ /^FEATURES\s+.*$/ || $thisline =~ /^FH\s+.*$/) {
# beginning of feature table, flags already set up
} elsif ($thisline =~ /^ORIGIN\s*.*$/ || $thisline =~ /^SQ\s*.*$/) {
# end of feature table, print final newline
flushline ();
if ($in_seq == 0) {
if ($organism ne "") {
print $FSA_OUT " [organism=$organism]";
$organism_ok = 1;
}
if ($topology ne "") {
print $FSA_OUT " [topology=$topology]";
}
if ($transl_table > 1) {
print $FSA_OUT " [gcode=$transl_table]";
}
print $FSA_OUT "\n";
}
$in_feat = 0;
$in_key = 0;
$in_qual = 0;
$is_source = 0;
$is_translation = 0;
$in_seq = 1;
} elsif ($thisline =~ /^\/\/\.*/) {
if ($organism_ok == 0) {
print $ERR_OUT "ERROR - No organism found\n";
}
# at end-of-record double slash, reset variables for catenated flatfiles
clearflags ();
} elsif ($in_seq == 1) {
if ($thisline =~ /^\s+\d+ (.*)$/ || $thisline =~ /^\s+(.*)\s+\d+$/) {
# report sequence
$curr_seq = $1;
$curr_seq =~ s/ //g;
$curr_seq = uc $curr_seq;
print $FSA_OUT "$curr_seq\n";
}
} elsif ($in_feat == 1) {
if ($thisline =~ /^ {1,10}(\w+)\s+(.*)$/ || $thisline =~ /^FT (\w+)\s+(.*)$/) {
# new feature key and location
flushline ();
$in_key = 1;
$in_qual = 0;
$current_key = $1;
$current_loc = $2;
if ($current_key =~ /source/ || $current_key =~ /Source/) {
$is_source = 1;
} else {
$is_source = 0;
}
} elsif ($thisline =~ /^\s+\/(\w+)=(.*)$/ || $thisline =~ /^FT\s+\/(\w+)=(.*)$/) {
# new qualifier
flushline ();
$in_key = 0;
$in_qual = 1;
$current_qual = $1;
# remove leading double quote
my $val = $2;
$val =~ s/\"//g;
$current_val = $val;
if ($current_qual =~ /translation/) {
$is_translation = 1;
} else {
$is_translation = 0;
}
if ($current_qual =~ /transl_table/) {
$transl_table = $current_val;
}
} elsif ($thisline =~ /^\s+\/(\w+)$/ || $thisline =~ /^FT\s+\/(\w+)$/) {
# new singleton qualifier - e.g., trans-splicing, pseudo
flushline ();
$in_key = 0;
$in_qual = 1;
$current_qual = $1;
$current_val = "";
$is_translation = 0;
} elsif ($thisline =~ /^\s+(.*)$/ || $thisline =~ /^FT\s+(.*)$/) {
if ($in_key == 1) {
# continuation of feature location
$current_loc = $current_loc . $1;
} elsif ($in_qual == 1) {
# continuation of qualifier
# remove trailing double quote
my $val = $1;
$val =~ s/\"//g;
if ($is_translation == 1) {
$current_val = $current_val . $val;
} else {
$current_val = $current_val . " " . $val;
}
}
}
}
}
# close input and output files
close ($GBF_IN);
close ($FSA_OUT);
close ($TBL_OUT);
close ($ERR_OUT);
# if no bad features or qualifiers, remove error file
if (! $has_errors) {
unlink ("$base.err");
}
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