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+ GNU AFFERO GENERAL PUBLIC LICENSE
+ Version 3, 19 November 2007
+
+ Copyright (C) 2007 Free Software Foundation, Inc. <https://fsf.org/>
+ Everyone is permitted to copy and distribute verbatim copies
+ of this license document, but changing it is not allowed.
+
+ Preamble
+
+ The GNU Affero General Public License is a free, copyleft license for
+software and other kinds of works, specifically designed to ensure
+cooperation with the community in the case of network server software.
+
+ The licenses for most software and other practical works are designed
+to take away your freedom to share and change the works. By contrast,
+our General Public Licenses are intended to guarantee your freedom to
+share and change all versions of a program--to make sure it remains free
+software for all its users.
+
+ When we speak of free software, we are referring to freedom, not
+price. Our General Public Licenses are designed to make sure that you
+have the freedom to distribute copies of free software (and charge for
+them if you wish), that you receive source code or can get it if you
+want it, that you can change the software or use pieces of it in new
+free programs, and that you know you can do these things.
+
+ Developers that use our General Public Licenses protect your rights
+with two steps: (1) assert copyright on the software, and (2) offer
+you this License which gives you legal permission to copy, distribute
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+
+ A secondary benefit of defending all users' freedom is that
+improvements made in alternate versions of the program, if they
+receive widespread use, become available for other developers to
+incorporate. Many developers of free software are heartened and
+encouraged by the resulting cooperation. However, in the case of
+software used on network servers, this result may fail to come about.
+The GNU General Public License permits making a modified version and
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+source code to the public.
+
+ The GNU Affero General Public License is designed specifically to
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+to the community. It requires the operator of a network server to
+provide the source code of the modified version running there to the
+users of that server. Therefore, public use of a modified version, on
+a publicly accessible server, gives the public access to the source
+code of the modified version.
+
+ An older license, called the Affero General Public License and
+published by Affero, was designed to accomplish similar goals. This is
+a different license, not a version of the Affero GPL, but Affero has
+released a new version of the Affero GPL which permits relicensing under
+this license.
+
+ The precise terms and conditions for copying, distribution and
+modification follow.
+
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+reviewing courts shall apply local law that most closely approximates
+an absolute waiver of all civil liability in connection with the
+Program, unless a warranty or assumption of liability accompanies a
+copy of the Program in return for a fee.
+
+ END OF TERMS AND CONDITIONS
+
+ How to Apply These Terms to Your New Programs
+
+ If you develop a new program, and you want it to be of the greatest
+possible use to the public, the best way to achieve this is to make it
+free software which everyone can redistribute and change under these terms.
+
+ To do so, attach the following notices to the program. It is safest
+to attach them to the start of each source file to most effectively
+state the exclusion of warranty; and each file should have at least
+the "copyright" line and a pointer to where the full notice is found.
+
+ <one line to give the program's name and a brief idea of what it does.>
+ Copyright (C) <year> <name of author>
+
+ This program is free software: you can redistribute it and/or modify
+ it under the terms of the GNU Affero General Public License as published by
+ the Free Software Foundation, either version 3 of the License, or
+ (at your option) any later version.
+
+ This program is distributed in the hope that it will be useful,
+ but WITHOUT ANY WARRANTY; without even the implied warranty of
+ MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE. See the
+ GNU Affero General Public License for more details.
+
+ You should have received a copy of the GNU Affero General Public License
+ along with this program. If not, see <https://www.gnu.org/licenses/>.
+
+Also add information on how to contact you by electronic and paper mail.
+
+ If your software can interact with users remotely through a computer
+network, you should also make sure that it provides a way for users to
+get its source. For example, if your program is a web application, its
+interface could display a "Source" link that leads users to an archive
+of the code. There are many ways you could offer source, and different
+solutions will be better for different programs; see section 13 for the
+specific requirements.
+
+ You should also get your employer (if you work as a programmer) or school,
+if any, to sign a "copyright disclaimer" for the program, if necessary.
+For more information on this, and how to apply and follow the GNU AGPL, see
+<https://www.gnu.org/licenses/>.
diff --git a/README.md b/README.md
index 47e9c45a802bb09fad4e0003af2a85ab224694a6..e2d5b678283901156773dd931098239b99026e84 100644
--- a/README.md
+++ b/README.md
@@ -1,3 +1,163 @@
# eCDS
-Extracting CDS from contig sequences
\ No newline at end of file
+_eCDS_ is a command line tool written in [Bash](https://www.gnu.org/software/bash/) to ease the extraction of coding sequence(s) (CDS) from a nucleotide sequence FASTA file (typically, a set of contig sequences). Given a specified query file (containing either an amino acid sequence or a position-specific scoring matrix), _eCDS_ first runs a TBLASTN similarity search (e.g. Gertz et al. 2006) against the specified subject sequence(s), and next extracts the CDS corresponding to each selected BLAST hit.
+
+_eCDS_ runs on UNIX, Linux and most OS X operating systems.
+
+
+
+
+## Dependencies
+
+You will need to install the required programs and tools listed in the following table, or to verify that they are already installed with the required version.
+
+<div align="center">
+
+| program | package | version | sources |
+|:----------------------------------------------------------------------------------- |:----------------------------------------------------------------:| ----------:|:--------------------------------------------------------------------------------------------------------- |
+| [_eFASTA_](https://gitlab.pasteur.fr/GIPhy/eFASTA) | - | ≥ 1.2b | [gitlab.pasteur.fr/GIPhy/eFASTA](https://gitlab.pasteur.fr/GIPhy/eFASTA) |
+| [_gawk_](https://www.gnu.org/software/gawk/) | - | > 4.0.0 | [ftp.gnu.org/gnu/gawk](http://ftp.gnu.org/gnu/gawk/) |
+| _makeblastdb_ <br> _tblastn_ | [blast+](https://bmcbioinformatics.biomedcentral.com/articles/10.1186/1471-2105-10-421) | ≥ 2.9.0 | [ftp.ncbi.nlm.nih.gov/blast/executables/blast+/LATEST](ftp://ftp.ncbi.nlm.nih.gov/blast/executables/blast+/LATEST) |
+
+</div>
+
+## Installation and execution
+
+**A.** Clone this repository with the following command line:
+
+```bash
+git clone https://gitlab.pasteur.fr/GIPhy/eCDS.git
+```
+
+**B.** Give the execute permission to the file `eCDS.sh`:
+
+```bash
+chmod +x eCDS.sh
+```
+
+**C.** Execute _eCDS_ with the following command line model:
+
+```bash
+./eCDS.sh [options]
+```
+
+**D.** If at least one of the required program (see [Dependencies](#dependencies)) is not available on your `$PATH` variable (or if one compiled binary has a different default name), _eCDS_ will exit with an error message.
+When running _eCDS_ without option, a documentation should be displayed; otherwise, the name of the missing program is displayed before existing.
+In such a case, edit the file `eCDS.sh` and indicate the local path to the corresponding binary(ies) within the code block `REQUIREMENTS` (approximately lines 50-90).
+For each required program, the table below reports the corresponding variable assignment instruction to edit (if needed) within the code block `REQUIREMENTS`
+
+<div align="center">
+<sup>
+
+| program | variable assignment | | program | variable assignment |
+|:---------------- |:-------------------------------- | - |:---------------- |:-------------------------------- |
+| _eFASTA_ | `EFASTA_BIN=eFASTA;` | | _makeblastdb_ | `MAKEBLASTDB_BIN=makeblastdb;` |
+| _gawk_ | `GAWK_BIN=gawk; ` | | _tblastn_ | `TBLASTN_BIN=tblastn;` |
+
+</sup>
+</div>
+
+Note that depending on the installation of some required programs, the corresponding variable can be assigned with complex commands.
+For example, as _eFASTA_ is a Java tool that can be run using a Java virtual machine, the executable jar file `EFASTA.jar` can be used by _eCDS_ by editing the corresponding variable assignment instruction as follows: `EFASTA_BIN="java -jar EFASTA.jar"`.
+
+
+## Usage
+
+Run _eCDS_ without option to read the following documentation:
+
+```
+ USAGE : eCDS -q <infile> -s <infile> -o <basename>
+ [-p <ppos>] [-c <qcov>] [-b <bitscore>]
+ [-S <integer>] [-f] [-a] [-h]
+
+ where options are :
+
+ -q <infile> query file containing either a sequence (FASTA format) or a position-
+ specific scoring matrix (PSSM; ASN.1 format)
+ -s <infile> subject sequence file (FASTA format)
+ -o <basename> to write the inferred CDS into the output file names [basename].fna
+ (codon) and [basename].faa (translated amino acids)
+ -p <integer> minimum allowed percentage of positive-scoring amino acid matches
+ (default: 50)
+ -c <integer> minimum allowed percentage of query coverage (default: 70)
+ -b <integer> minimum allowed bit score value (default: 0)
+ -f writes only CDS inferred from the first best hit (default: not set)
+ -S <integer> to accept alternate START codons when searching for the complete CDS:
+ 1 = ATG (default)
+ 2 = ATG GTG TTG
+ 3 = ATG GTG TTG CTG ATT
+ 4 = ATG GTG TTG CTG ATT ATC ATA
+ -a appends CDS to the end of the FASTA output files when they already
+ exist (default: content replaced)
+ -h prints this help and exits
+```
+
+## Notes
+
+* _eCDS_ is able to consider as query (option `-q`) either a single amino acid sequence in FASTA format or a position-specific scoring matrix (PSSM) in ASN.1 format. When using a FASTA file as query, only the first sequence is considered. When using a PSSM file as query, rescaling is automatically performed to be read by the _tblastn_ binary (i.e. scalingFactor = 1 into the ASN.1-formatted PSSM file).
+
+* A PSSM file summarizes a multiple amino acid alignment; it is therefore very useful to use a PSSM as a generic query against genome sequences belonging to any genera. A very large collection of PSSM can be found at [ftp.ncbi.nih.gov/pub/mmdb/cdd/](https://ftp.ncbi.nih.gov/pub/mmdb/cdd/) (into the archive [_cdd.tar.gz_](https://ftp.ncbi.nih.gov/pub/mmdb/cdd/cdd.tar.gz)). For a given gene name (e.g. "ribosomal protein L3"), the associated conserved domain PSSM identifier(s) can be easily retrieved using the search tool at [www.ncbi.nlm.nih.gov/cdd](https://www.ncbi.nlm.nih.gov/cdd/) or [www.ncbi.nlm.nih.gov/proteinclusters](https://www.ncbi.nlm.nih.gov/proteinclusters); complete lists are also available at [ftp.ncbi.nih.gov/genomes/CLUSTERS/](https://ftp.ncbi.nih.gov/genomes/CLUSTERS/) (files _\*\_clusters.txt_). Moreover, for a given CDS with GenBank identifier (e.g. NP_417779), the associated conserved domain PSSM identifier(s) can be retrieved using the following URL: [www.ncbi.nlm.nih.gov/Structure/cdd/wrpsb.cgi?FULL&SEQUENCE=NP_417779](https://www.ncbi.nlm.nih.gov/Structure/cdd/wrpsb.cgi?FULL&SEQUENCE=NP_417779) (juste change the accession identifier at the end of the URL). Note that using PSSM generated from the automatically aligned sequences classified as stable clusters in the [NCBI Protein Clusters database](https://www.ncbi.nlm.nih.gov/proteinclusters) (i.e. PRK data) enables to observe accurate results when using _eCDS_.
+
+* After carrying out a TBLASTN search of the specified query (option `-q`) against the specified subject FASTA file (option `-s`), non-significant BLAST hits are filtered out according three possible criteria: <br> • minimum percentage of positive-scoring amino matches (option `-p`; default: ppos = 50\%), <br> • minimum percentage of query coverage (option `-c`; default: qcov = 70\%), <br> • minimum bit score value (option `-b`; default: bitscore = 0). <br> The first criterion cutoff can be enlarged when using a query sequence that is expected to be closely related to the subject sequence (e.g. `-p 80`), as well as the second criterion cutoff. The third criterion can be useful when using one of the query PSSM distributed by the NCBI, as a minimum bitscore threshold is available for each of them. For example, PRK05644, a PSSM associated to the gene _gyrB_, can be used as query with the bitscore threshold specified at the corresponding webpage: [www.ncbi.nlm.nih.gov/Structure/cdd/cddsrv.cgi?uid=PRK05644](https://www.ncbi.nlm.nih.gov/Structure/cdd/cddsrv.cgi?uid=PRK05644) ('+ Statistics' on the left); note also that updated lists of bitscore thresholds are available at [ftp.ncbi.nih.gov/pub/mmdb/cdd/](https://ftp.ncbi.nih.gov/pub/mmdb/cdd/) (files _bitscore\_specific\_\*.txt_). However, a simple approach is also to run _eCDS_ with relaxed criteria and only consider the first best BLAST hit (option `-f`).
+
+* Finally, for each selected BLAST hit, the corresponding subject sequence region is considered to extract the smallest putative CDS containing that region, using [_eFASTA_](https://gitlab.pasteur.fr/GIPhy/eFASTA). Note that the determination of the CDS can be tuned by accepting alternate START codons (option `-S`).
+
+* Amino acid and codon sequences are written into FASTA files with extensions .faa and .fna, respectively. The base name the the output files should be specified using option `-o`. Note that a log file containing several BLAST results is also written (extension .log.txt).
+
+
+
+## Example
+
+In order to illustrate the usefulness of _eCDS_, the directory _example/_ contains the whole chromosome sequence of _Bacillus subtilis_ strain 168<sup>T</sup> (GenBank accession [AL009126.3](https://www.ncbi.nlm.nih.gov/nuccore/AL009126.3/)), as well as four PSSM files:
+<br> • _PRK00013.groEL.smp_ corresponding to the conserved domain [PRK00013](https://www.ncbi.nlm.nih.gov/Structure/cdd/cddsrv.cgi?uid=PRK00013) associated to the gene _groEL_,
+<br> • _PRK00380 .panC.smp_ corresponding to the conserved domain [PRK00380](https://www.ncbi.nlm.nih.gov/Structure/cdd/cddsrv.cgi?uid=PRK00380) associated to the gene _panC_,
+<br> • _PRK00405.rpoB.smp_ corresponding to the conserved domain [PRK00405](https://www.ncbi.nlm.nih.gov/Structure/cdd/cddsrv.cgi?uid=PRK00405) associated to the gene _rpoB_,
+<br> • _PRK05644.gyrB.smp_ corresponding to the conserved domain [PRK05644](https://www.ncbi.nlm.nih.gov/Structure/cdd/cddsrv.cgi?uid=PRK05644) associated to the gene _gyrB_.
+
+
+Basically, the four following command lines can be used to extract the four _groEL_, _panC_, _rpoB_ and _gyrB_ CDS from the _B. subtilis_ 168<sup>T</sup> chromosome:
+```bash
+eCDS.sh -q PRK00013.groEL.smp -s Bacillus.subtilis.168.fasta -o bc.groEL
+eCDS.sh -q PRK00380.panC.smp -s Bacillus.subtilis.168.fasta -o bc.panC
+eCDS.sh -q PRK00405.rpoB.smp -s Bacillus.subtilis.168.fasta -o bc.rpoB
+eCDS.sh -q PRK05644.gyrB.smp -s Bacillus.subtilis.168.fasta -o bc.gyrB
+```
+
+For _groEL_ gene, the output file _bc.groEL.faa_ (see _example/_) contains the expected amino acid sequence, i.e. identical to the annotated _groEL_ gene within the _B. subtilis_ 168<sup>T</sup> chromosome [AL009126.3](https://www.ncbi.nlm.nih.gov/nuccore/AL009126.3/) (i.e. [CAB12422](https://www.ncbi.nlm.nih.gov/protein/2632916)). For _panC_ gene, _eCDS_ also extracts an amino acid sequence (_bc.panC.faa_ in _example/_) that is identical to the annotated one (i.e. [CAB14158](https://www.ncbi.nlm.nih.gov/protein/2634660)).
+
+For _rpoB_ gene, one can observe that the above command line leads to an amino acid sequence that is not starting with a START codon:
+
+```
+>AL009126.3::121907-125497
+GVNQLTGQLVQYGRHRQRRSYARISEVLELPNLIEIQTSSYQWFLDEGLREMFQDISPIEDFTGNLSLEFIDYSLGEPKYPVEESKERDVTYSAPLRVKVRLINKETGEVKDQDVFMGDFPIMTDTGTFIINGAERVIVSQLVRSPSVYFSGKVDKNGKKGFTATVIPNRGAWLEYETDAKDVVYVRIDRTRKLPVTVLLRALGFGSDQEILDLIGENEYLRNTLDKDNTENSDKALLEIYERLRPGEPPTVENAKSLLDSRFFDPKRYDLANVGRYKINKKLHIKNRLFNQRLAETLVDPETGEILAEKGQILDRRTLDKVLPYLENGIGFRKLYPNGGVVEDEVTLQSIKIFAPTDQEGEQVINVIGNAYIEEEIKNITPADIISSISYFFNLLHGVGDTDDIDHLGNRRLRSVGELLQNQFRIGLSRMERVVRERMSIQDTNTITPQQLINIRPVIASIKEFFGSSQLSQFMDQTNPLAELTHKRRLSALGPGGLTRERAGMEVRDVHYSHYGRMCPIETPEGPNIGLINSLSSYAKVNRFGFIETPYRRVDPETGKVTGRIDYLTADEEDNYVVAQANARLDDEGAFIDDSIVARFRGENTVVSRNRVDYMDVSPKQVVSAATACIPFLENDDSNRALMGANMQRQAVPLMQPEAPFVGTGMEYVSGKDSGAAVICKHPGIVERVEAKNVWVRRYEEVDGQKVKGNLDKYSLLKFVRSNQGTCYNQRPIVSVGDEVVKGEILADGPSMELGELALGRNVMVGFMTWDGYNYEDAIIMSERLVKDDVYTSIHIEEYESEARDTKLGPEEITRDIPNVGEDALRNLDDRGIIRIGAEVKDGDLLVGKVTPKGVTELTAEERLLHAIFGEKAREVRDTSLRVPHGGGGIIHDVKVFNREDGDELPPGVNQLVRVYIVQKRKISEGDKMAGRHGNKGVISKILPEEDMPYLPDGTPIDIMLNPLGVPSRMNIGQVLELHMGMAARYLGIHIASPVFDGAREEDVWETLEEAGMSRDAKTVLYDGRTGEPFDNRVSVGIMYMIKLAHMVDDKLHARSTGPYSLVTQQPLGGKAQFGGQRFGEMEVWALEAYGAAYTLQEILTVKSDDVVGRVKTYEAIVKGDNVPEPGVPESFKVLIKELQSLGMDVKILSGDEEEIEMRDLEDEEDAKQADGLALSGDEEPEETASADVERDVVTKE
+```
+
+However, such a problem can be generally fixed by using alternate START codons. This can be achieved by the following example command line:
+
+```bash
+eCDS.sh -q PRK00405.rpoB.smp -s Bacillus.subtilis.168.fasta -S 2 -o bc.rpoB
+```
+
+that writes the output file _bc.rpoB.faa_ (see _example/_) containing an amino acid sequence that is identical to the annotated _B. subtilis_ 168<sup>T</sup> _rpoB_ CDS (i.e. [CAB11883](https://www.ncbi.nlm.nih.gov/protein/225184649)).
+
+Finally, for _gyrB_ gene, one can observe that the above command line leads to two extracted sequences. As expected, the first one (i.e. first best BLAST hit) is identical to the _B. subtilis_ 168<sup>T</sup> _gyrB_ CDS (i.e. [CAB11782](https://www.ncbi.nlm.nih.gov/protein/2632273)). However, the second sequence corresponds to the gene _parE_ (subunit B of DNA topoisomerase IV), a well-known paralog of _gyrB_ (e.g. Poirier et al. 2018). A way to circumvent this problem is to only consider the first best BLAST hit (option `-f`). However, an alternative way is to use the bitscore threshold suggested for PRK05644, available at [www.ncbi.nlm.nih.gov/Structure/cdd/cddsrv.cgi?uid=PRK05644](https://www.ncbi.nlm.nih.gov/Structure/cdd/cddsrv.cgi?uid=PRK05644); this leads to the following example command line:
+
+```bash
+eCDS.sh -q PRK05644.gyrB.smp -s Bacillus.subtilis.168.fasta -p 0 -c 0 -b 1029 -o bc.gyrB
+```
+
+that writes only the _gyrB_ amino acid and codon sequences into the files _bc.rpoB.faa_ and _bc.rpoB.fna_, respectively (see _example/_).
+
+
+
+
+
+
+## References
+
+Gertz EM, Yu YK, Agarwala R, Schäffer AA, Altschul SF (2006) _Composition-based statistics and translated nucleotide searches: Improving the TBLASTN module of BLAST_. **BMC Biology**, 4:41. [doi:10.1186/1741-7007-4-41](https://doi.org/10.1186/1741-7007-4-41).
+
+Poirier S, Rué O, Peguilhan R, Coeuret G, Zagorec M, Champomier-Vergès M-C, Loux V, Chaillou S (2018) _Deciphering intra-species bacterial diversity of meat and seafood spoilage microbiota using gyrB amplicon sequencing: A comparative analysis with 16S rDNA V3-V4 amplicon sequencing_. **PLoS One**, 13(9):e0204629. [doi:10.1371/journal.pone.0204629](https://doi.org/10.1371/journal.pone.0204629).
+
+
+
diff --git a/eCDS.sh b/eCDS.sh
new file mode 100755
index 0000000000000000000000000000000000000000..b1b65fd8b5b177b3f75a39a43d8f8e43c7108dd1
--- /dev/null
+++ b/eCDS.sh
@@ -0,0 +1,297 @@
+#!/bin/bash
+
+##############################################################################################################
+# #
+# eCDS: extracting CDS segment from contig sequences #
+# #
+ COPYRIGHT="Copyright (C) 2016-2021 Institut Pasteur" #
+# #
+# This program is free software: you can redistribute it and/or modify it under the terms of the GNU #
+# General Public License as published by the Free Software Foundation, either version 3 of the License, or #
+# (at your option) any later version. #
+# #
+# This program is distributed in the hope that it will be useful, but WITHOUT ANY WARRANTY; without even #
+# the implied warranty of MERCHANTABILITY or FITNESS FOR A PARTICULAR PURPOSE. See the GNU General Public #
+# License for more details. #
+# #
+# You should have received a copy of the GNU General Public License along with this program. If not, see #
+# <http://www.gnu.org/licenses/>. #
+# #
+# Contact: #
+# Alexis Criscuolo alexis.criscuolo@pasteur.fr #
+# Genome Informatics & Phylogenetics (GIPhy) giphy.pasteur.fr #
+# Bioinformatics and Biostatistics Hub research.pasteur.fr/en/team/hub-giphy #
+# Dpt. Biologie Computationnelle research.pasteur.fr/team/bioinformatics-and-biostatistics-hub #
+# Institut Pasteur, Paris, FRANCE research.pasteur.fr #
+# #
+##############################################################################################################
+
+##############################################################################################################
+# #
+# ============ #
+# = VERSIONS = #
+# ============ #
+# #
+ VERSION=1.0.210724ac; #
+# #
+##############################################################################################################
+
+##############################################################################################################
+# #
+# ============= #
+# = CONSTANTS = #
+# ============= #
+# #
+# -- PWD: directory containing the current script --------------------------------------------------------- #
+# #
+ PWD="$(cd "$(dirname "${BASH_SOURCE[0]}")" >/dev/null 2>&1 && pwd)";
+# #
+# -- constants -------------------------------------------------------------------------------------------- #
+# #
+ NA="._N.o.N._.A.p.P.l.I.c.A.b.L.e_.";
+# #
+##############################################################################################################
+
+##############################################################################################################
+# #
+# ================ #
+# = REQUIREMENTS = #
+# ================ #
+# #
+# -- gawk version >= 4 ------------------------------------------------------------------------------------ #
+# #
+ GAWK_BIN=gawk;
+ if [ ! $(command -v $GAWK_BIN) ]; then echo "$GAWK_BIN not found" >&2 ; exit 1 ; fi
+ BAWK="$GAWK_BIN";
+ CAWK="$GAWK_BIN -F,";
+ TAWK="$GAWK_BIN -F\\t";
+# #
+# -- makeblastdb & tblastn from NCBI blast+ package----- -------------------------------------------------- #
+# #
+ MAKEBLASTDB_BIN=makeblastdb;
+ if [ ! $(command -v $MAKEBLASTDB_BIN) ]; then echo "$MAKEBLASTDB_BIN not found" >&2 ; exit 1 ; fi
+ MAKEBLASTDB_STATIC_OPTIONS="-dbtype nucl";
+ MAKEBLASTDB="$MAKEBLASTDB_BIN $MAKEBLASTDB_STATIC_OPTIONS";
+ TBLASTN_BIN=tblastn;
+ if [ ! $(command -v $TBLASTN_BIN) ]; then echo "$TBLASTN_BIN not found" >&2 ; exit 1 ; fi
+ TBLASTN_STATIC_OPTIONS="-seg no -word_size 2 -xdrop_gap_final 1000";
+ TBLASTN="$TBLASTN_BIN $TBLASTN_STATIC_OPTIONS";
+# #
+# -- makeblastdb & tblastn from NCBI blast+ package----- -------------------------------------------------- #
+# #
+ EFASTA_BIN=eFASTA;
+ if [ ! $(command -v $EFASTA_BIN) ]; then echo "$EFASTA_BIN not found" >&2 ; exit 1 ; fi
+ EFASTA_STATIC_OPTIONS="-fcds";
+ EFASTA="$EFASTA_BIN $EFASTA_STATIC_OPTIONS";
+# #
+##############################################################################################################
+
+
+##############################################################################################################
+# #
+# ============ #
+# = DOC = #
+# ============ #
+# #
+mandoc() {
+ echo -e "\n\033[1m eCDS v$VERSION $COPYRIGHT\033[0m";
+ cat <<EOF
+
+ https://gitlab.pasteur.fr/GIPhy/eCDS
+
+ USAGE : eCDS -q <infile> -s <infile> -o <basename>
+ [-p <ppos>] [-c <qcov>] [-b <bitscore>]
+ [-S <integer>] [-f] [-a] [-h]
+
+ where options are :
+
+ -q <infile> query file containing either a sequence (FASTA format) or a position-
+ specific scoring matrix (PSSM; ASN.1 format)
+ -s <infile> subject sequence file (FASTA format)
+ -o <basename> to write the inferred CDS into the output file names [basename].fna
+ (codon) and [basename].faa (translated amino acids)
+ -p <integer> minimum allowed percentage of positive-scoring amino acid matches
+ (default: 50)
+ -c <integer> minimum allowed percentage of query coverage (default: 70)
+ -b <integer> minimum allowed bit score value (default: 0)
+ -f writes only CDS inferred from the first best hit (default: not set)
+ -S <integer> to accept alternate START codons when searching for the complete CDS:
+ 1 = ATG (default)
+ 2 = ATG GTG TTG
+ 3 = ATG GTG TTG CTG ATT
+ 4 = ATG GTG TTG CTG ATT ATC ATA
+ -a appends CDS to the end of the FASTA output files when they already
+ exist (default: content replaced)
+ -h prints this help and exits
+
+EOF
+}
+# #
+##############################################################################################################
+
+##############################################################################################################
+# #
+# ============= #
+# = FUNCTIONS = #
+# ============= #
+# #
+# -- runcmd ----------------------------------------------------------------------------------------------- #
+# >> executes the specified command line $1; exit 1 if the command line returns an error signal #
+# #
+runcmd () {
+ sh -c "$*" || { echo "FAIL: $*" ; exit 1 ; } ;
+}
+# #
+# -- randfile --------------------------------------------------------------------------------------------- #
+# >> creates and returns a random file name that does not exist from the specified basename $1 #
+# #
+randfile() {
+ local rdf="$(mktemp $1.XXXXXXXXX)";
+ echo $rdf ;
+}
+# #
+##############################################################################################################
+
+
+##############################################################################################################
+# #
+# READING OPTIONS #
+# #
+##############################################################################################################
+
+if [ $# -lt 1 ]; then mandoc ; exit 1 ; fi
+
+QUERY="$NA";
+SUBJECT="$NA";
+BASEFILE="$NA";
+MINPPOS=50;
+MINQCOV=70;
+MINBITS=0;
+CSTART=1;
+APPEND=false;
+ONLYFIRST=false;
+
+while getopts :q:s:o:p:c:b:S:afh option
+do
+ case $option in
+ q) QUERY=$OPTARG ;;
+ s) SUBJECT=$OPTARG ;;
+ o) BASEFILE=$OPTARG ;;
+ p) MINPPOS=$OPTARG ;;
+ c) MINQCOV=$OPTARG ;;
+ b) MINBITS=$OPTARG ;;
+ S) CSTART=$OPTARG ;;
+ a) APPEND=true ;;
+ f) ONLYFIRST=true ;;
+ h) mandoc ; exit 0 ;;
+ \?) mandoc ; exit 1 ;;
+ esac
+done
+
+if [ "$QUERY" == "$NA" ]; then echo "no query file (option -q)" >&2 ; exit 1 ; fi
+if [ "$SUBJECT" == "$NA" ]; then echo "no subject file (option -s)" >&2 ; exit 1 ; fi
+if [ "$BASEFILE" == "$NA" ]; then echo "no basename specified (option -o)" >&2 ; exit 1 ; fi
+if [ ! -s $QUERY ]; then echo "no or empty query file (option -q): $QUERY" >&2 ; exit 1 ; fi
+if [ ! -s $SUBJECT ]; then echo "no or empty subject file (option -s): $INFILE" >&2 ; exit 1 ; fi
+if ! [[ $MINPPOS =~ ^[0-9]+$ ]]; then echo "incorrect integer value (option -p): $MINPPOS" >&2 ; exit 1 ; fi
+if [ $MINPPOS -gt 100 ]; then echo "integer percentage too high (option -p): $MINPPOS" >&2 ; exit 1 ; fi
+if ! [[ $MINQCOV =~ ^[0-9]+$ ]]; then echo "incorrect integer value (option -c): $MINQCOV" >&2 ; exit 1 ; fi
+if [ $MINQCOV -gt 100 ]; then echo "integer percentage too high (option -c): $MINQCOV" >&2 ; exit 1 ; fi
+if ! [[ $MINBITS =~ ^[0-9]+$ ]]; then echo "incorrect integer value (option -b): $MINBITS" >&2 ; exit 1 ; fi
+
+
+##############################################################################################################
+# #
+# GO!! #
+# #
+##############################################################################################################
+
+export LC_ALL=C ;
+
+TBNOUT=$BASEFILE.tbn ;
+LOG=$BASEFILE.log.txt ;
+TMP=$(randfile $BASEFILE);
+QRY=$(randfile $QUERY);
+
+trap "echo -n interrupting ; echo -n . ; kill -9 $(jobs -pr) &> /dev/null ; echo -n . ; rm -f $TBNOUT $TMP $QRY $SUBJECT.{ndb,nhr,nin,not,nsq,ntf,nto} &>/dev/null ; echo -n . ; wait ; exit 1 " SIGINT
+
+if ! $APPEND
+then
+ [ -e $BASEFILE.faa ] && rm -f $BASEFILE.faa ;
+ [ -e $BASEFILE.fna ] && rm -f $BASEFILE.fna ;
+fi
+[ "$CSTART" == "2" ] && EFASTA="$EFASTA -start PROK";
+[ "$CSTART" == "3" ] && EFASTA="$EFASTA -start PROK+";
+[ "$CSTART" == "4" ] && EFASTA="$EFASTA -start PROK++";
+
+echo -n -e "SUBJECT: $(basename $SUBJECT)\t\t";
+
+$MAKEBLASTDB -in $SUBJECT > /dev/null ;
+
+echo -n -e "QUERY: $(basename $QUERY)\t\t";
+
+if [ $(grep -c -m1 "^>" $QUERY) -eq 0 ]
+then
+ optin="-in_pssm";
+ # rescaling PSSM
+ sf=$(grep "scalingFactor" $QUERY | $BAWK '{print$2}' | tr -d ',');
+ $BAWK -v sf=$sf '($1=="scores") {rescale=1;print;next}
+ (rescale&&$1=="},") {rescale=0}
+ (rescale) {s=$1;n=sub(",","",s);s=sprintf("%d",(s/sf));printf" "s;print (n==0)?"":",";next}
+ ($1=="scalingFactor"){print" scalingFactor 1,";next}
+ {print}' $QUERY > $QRY ;
+else
+ optin="-query";
+ # getting the first sequence
+ $BAWK '!/^>/{s=s$0;next}(s!=""){print s;s=""}{print}END{print s}' $QUERY | head -2 > $QRY ;
+fi
+
+
+echo -e "#query\tsubject\tsstart\tsend\tpident\tppos\tqcov\tevalue\tbitscore" > $TBNOUT ;
+# fields= 1 2 3 4 5 6 7 8 9 10
+$TBLASTN $optin $QRY -db $SUBJECT -outfmt "6 saccver qlen qstart qend sstart send pident ppos evalue bitscore" |
+ $TAWK -v q=$(basename ${QUERY%.*}) -v p=$MINPPOS -v c=$MINQCOV -v b=$MINBITS '($10>=b&&$8>=p&&(qcov=(100*($4-$3+1)/$2))>=c){print q"\t"$1"\t"$5"\t"$6"\t"$7"\t"$8"\t"qcov"\t"$9"\t"$10}' >> $TBNOUT ;
+
+rm -f $SUBJECT.{ndb,nhr,nin,not,nsq,ntf,nto} $QRY ;
+
+n=$(grep -v "^#query" $TBNOUT | wc -l);
+if [ $n -eq 0 ]
+then
+ echo "no hit found" ;
+ rm -f $TBNOUT $TMP ;
+ exit ;
+fi
+if [ ! -s $LOG ]
+then
+ cat $TBNOUT >> $LOG ;
+else
+ grep -v "^#query" $TBNOUT >> $LOG ;
+fi
+if [ $n -eq 1 ]
+then
+ echo -n "one hit found" ;
+else
+ echo -n "$n hits found" ;
+fi
+
+while read -r _ id s e _ _ _ _ _
+do
+ if [ "$s" == "sstart" ]; then continue ; fi
+ $EFASTA -fasta $SUBJECT -coord $id:$s-$e -outname $TMP ;
+ cat $TMP.faa >> $BASEFILE.faa ;
+ cat $TMP.fna >> $BASEFILE.fna ;
+ rm -f $TMP.faa $TMP.fna ;
+ if $ONLYFIRST ; then break ; fi
+done < $TBNOUT
+
+if [ $n -eq 1 ]
+then
+ lgt=$(sed -n '2p' $BASEFILE.faa | tr -d '\n' | wc -c);
+ echo -n " ($lgt aa)" ;
+fi
+echo ": $(basename $BASEFILE).faa $(basename $BASEFILE).fna" ;
+
+rm -f $TBNOUT $TMP ;
+
+exit
+
diff --git a/example/Bacillus.subtilis.168.fasta b/example/Bacillus.subtilis.168.fasta
new file mode 100644
index 0000000000000000000000000000000000000000..6292bec99563fddbd3edef172cfefb996aff09e2
--- /dev/null
+++ b/example/Bacillus.subtilis.168.fasta
@@ -0,0 +1,52697 @@
+>