LOCUS       Exported                7803 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Cloning vector pEAI3-S54, complete sequence.
ACCESSION   JQ733380
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7803)
  AUTHORS   Derouazi M, Toussaint B, Quenee L, Epaulard O, Guillaume M, Marlu R,
            Polack B.
  TITLE     High-yield production of secreted active proteins by the Pseudomonas
            aeruginosa type III secretion system
  JOURNAL   Appl. Environ. Microbiol. 74 (11), 3601-3604 (2008)
  PUBMED    18390679
REFERENCE   2  (bases 1 to 7803)
  AUTHORS   Toussaint B, Le Gouellec A, Polack B.
  TITLE     Direct Submission
  JOURNAL   Submitted (07-FEB-2012) TIMC-TheREx Laboratory, Universite Joseph 
            Fourier, Doamine de la merci, La Tronche 38700, France
REFERENCE   3  (bases 1 to 7803)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..7803
                     /organism="Cloning vector pEAI3-S54"
                     /lab_host="Pseudomonas aeruginosa strain CHA"
                     /mol_type="other DNA"
                     /PCR_primers="fwd_name: exsas, fwd_seq:
                     ccgaattcttataatatgcaaggag, fwd_name: xbaptaq, fwd_seq: 
                     cgtctagattgacaattcatcgcctcg, fwd_name: links, fwd_seq: 
                     gatccaccggtgctagcatgtacagagatctagcatg, fwd_name: 
                     orf1-exos54s, fwd_seq: catctagagggcaggtgctccggct, rev_name:
                     exsar, rev_seq: ggaagctttcaaaaaacgtcagtta, rev_name: 
                     kpnilaciq, rev_seq: cgggtacctcactgcccgctttccagt, rev_name: 
                     linkr, rev_seq: ctagatctctgtacatgctagcaccggtg, rev_name: 
                     orf1-exos54r, rev_seq: 
                     cttgcatgcctgcaggtcgacggatcctagcagcccctcaccct"
                     /db_xref="taxon:1157690"
     CDS             1..162
                     /codon_start=1
                     /gene="exoS"
                     /product="ExoS54"
                     /label=exoS
                     /note="secretion tag; first 54 amino acid of ExoS"
                     /protein_id="AFD22622.1"
                     /translation="MHIQSLQQSPSFAVELHQAASGRLGQIEARQVATPSEAQQLAQRQ
                     DAPKGEGLL"
     gene            1..162
                     /gene="exoS"
                     /label=exoS
     primer_bind     complement(211..227)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(388..1221)
                     /codon_start=1
                     /gene="repB"
                     /product="RepB"
                     /label=repB
                     /protein_id="AFD22623.1"
                     /translation="MASPPMVYKSNALVEAAYRLSVQEQRIVLACISQVKRSEPVTDEV
                     MYSVTAEDIATMAGVPIESSYNQLKEAALRLKRREVRLTQEPNGKGKRPSVMITGWVQT
                     IIYREGEGRVELRFTKDMLPYLTELTKQFTKYALADVAKMDSTHAIRLYELLMQWDSIG
                     QREIEIDQLRKWFQLEGRYPSIKDFKLRVLDPAVTQINEHSPLQVEWAQRKTGRKVTHL
                     LFSFGPKKPAKAVGKAPAKRKAGKISDAEIAKQARPGETWEAARARLTQMPLDLA"
     gene            complement(388..1221)
                     /gene="repB"
                     /label=repB
     rep_origin      1235..1586
                     /label=pRO1600 oriV
                     /note="broad-host-range origin of replication from 
                     Pseudomonas aeruginosa plasmid pRO1600; requires the 
                     pRO1600 Rep protein for replication (West et al., 1994)"
     promoter        1913..2017
                     /gene="bla"
                     /label=AmpR promoter
     CDS             2018..2878
                     /codon_start=1
                     /gene="bla"
                     /product="beta lactamase"
                     /label=bla
                     /protein_id="AFD22627.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            2018..2878
                     /gene="bla"
                     /label=bla
     rep_origin      3049..3637
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     protein_bind    3925..3946
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        3961..3991
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3999..4015
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     4023..4039
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(4067..5149)
                     /codon_start=1
                     /gene="lacIq"
                     /product="LacI"
                     /label=lacIq
                     /protein_id="AFD22624.1"
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGREHLVALGHQQIALLAGPLSSVSARLRLAGWHNFSLGNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     gene            complement(4067..5149)
                     /gene="lacIq"
                     /label=lacIq
     promoter        complement(5150..5227)
                     /gene="lacI (mutant)"
                     /label=lacIq promoter
                     /note="In the lacIq allele, a single base change in the 
                     promoter boosts expression of the lacI gene about 10-fold."
     terminator      5596..5623
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
     terminator      5716..5802
                     /gene="Escherichia coli rrnB"
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB 
                     gene"
     primer_bind     6203..6219
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(6237..7073)
                     /codon_start=1
                     /gene="exsA"
                     /product="ExsA"
                     /label=exsA
                     /protein_id="AFD22625.1"
                     /translation="MQGAKSLGRKQITSCHWNIPTFEYRVNKEEGVYVLLEGELTVQDI
                     DSTFCLAPGELLFVRRGSYVVSTKGKDSRILWIPLSAQFLQGFVQRFGALLSEVEHCDE
                     PVPGIIAFAATPLLAGCVKGLKELLVHEHPPMLACLKIEELLMLFAFSPQGPLLMSVLR
                     QLSNRHVERLQLFMEKHYLNEWKLSDFSREFGMGLTTFKELFGSVYGVSPRAWISERRI
                     LYAHQLLLNSDMSIVDIAMEAGFSSQSYFTQSYRRRFGCTPSRSRQGKDECRAKNN"
     gene            complement(6237..7073)
                     /gene="exsA"
                     /label=exsA
     protein_bind    7108..7124
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     CDS             complement(7266..7616)
                     /codon_start=1
                     /product="SpcS"
                     /label=SpcS
                     /protein_id="AFD22626.1"
                     /translation="MNPLYRAAIHQLFLALDLPTPNDEESVLSLQVGPHLCHLAEHPTD
                     HLLMFTRLEGQGDATASEQNLFSQDPCKPILGRDPESGERLLWNRQPLQLLDRAQIHHQ
                     LEQLVAAAEELR"