LOCUS       Exported                6744 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pFlagTEM1, complete sequence.
ACCESSION   EU440730
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6744)
  AUTHORS   Manuela R, Sun Y, Wilson PR, Tran QT, Chessa D, Andrews-Polymenis 
            HL, Lawhon SD, Figueiredo JF, Tsolis RM, Adams GL, Baumler AJ.
  TITLE     Host restriction of Salmonella enterica serotype Typhi is not caused
            by functional alteration of SipA, SopB, or SopD
  JOURNAL   Infect. Immun. 73 (12), 7817-7826 (2005)
  PUBMED    16299271
REFERENCE   2  (bases 1 to 6744)
  AUTHORS   Sun YH, Rolan HG, Tsolis RM.
  TITLE     Injection of flagellin into the host cell cytosol by Salmonella 
            enterica serotype Typhimurium
  JOURNAL   J. Biol. Chem. 282 (47), 33897-33901 (2007)
  PUBMED    17911114
REFERENCE   3  (bases 1 to 6744)
  AUTHORS   Sun Y, Tsolis RM.
  TITLE     Direct Submission
  JOURNAL   Submitted (31-JAN-2008) Medical Microbiology and Immunology, 
            UCDavis, School of Medicine, 451 Health Science Dr. GBSF Room 5415A,
            Davis, CA 95616, USA
REFERENCE   4  (bases 1 to 6744)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..6744
                     /organism="Cloning vector pFlagTEM1"
                     /mol_type="other DNA"
                     /db_xref="taxon:504831"
     misc_feature    join(1..1116,3531..6744)
                     /label=backbone from pBBR1MCS
                     /note="backbone from pBBR1MCS"
     promoter        371..473
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     misc_structure  1117..2590
                     /label=LacIq and Trc promoter from pTrc99A
                     /note="LacIq and Trc promoter from pTrc99A"
     promoter        1126..1203
                     /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."
     CDS             1204..2286
                     /codon_start=1
                     /gene="lacI"
                     /product="lac repressor"
                     /label=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)."
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        2518..2547
                     /label=trc promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     protein_bind    2555..2571
                     /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)."
     misc_feature    2603..2668
                     /label=3x Flag epitope
                     /note="3x Flag epitope"
     CDS             2603..2668
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     misc_feature    2675..3463
                     /label=TEM-1 without lead sequence
                     /note="TEM-1 without lead sequence"
     CDS             2678..3463
                     /codon_start=1
                     /gene="bla(M)"
                     /product="beta-lactamase lacking the signal sequence"
                     /label=bla(M)
                     /note="allows cytosolic expression of beta-lactamase"
                     /translation="PETLVKVKDAEDQLGARVGYIELDLNSGKILESFRPEERFPMMST
                     FKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYSPVTEKHLTDGMTVRELCSAAITMS
                     DNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRWEPELNEAIPNDERDTTMPVAMATT
                     LRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSALPAGWFIADKSGAGERGSRGIIAA
                     LGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGASLIKHW"
     promoter        complement(3539..3557)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3567..3583)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(4290..4952)
                     /codon_start=1
                     /product="replication protein for the broad-host-range 
                     plasmid pBBR1 from Bordetella bronchiseptica"
                     /label=pBBR1 Rep
                     /translation="MATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAAMLLH
                     HLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVSAYVV
                     NDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTGPGEE
                     PPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLPRR"
     rep_origin      4953..5722
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"