LOCUS       Exported                4503 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Expression vector pColdTEV(tet), complete sequence.
ACCESSION   FJ861704
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4503)
  AUTHORS   Schneider WM, Inouye M, Montelione GT, Roth MJ.
  TITLE     Independently inducible system of gene expression for condensed 
            single protein production (cSPP) suitable for high efficiency 
            isotope enrichment
  JOURNAL   J. Struct. Funct. Genomics 10 (3), 219-225 (2009)
  PUBMED    19642019
REFERENCE   2  (bases 1 to 4503)
  AUTHORS   Schneider WM, Roth MJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (23-MAR-2009) Biochemistry, UMDNJ, 675 Hoes Lane, 
            Piscataway, NJ 08854, USA
REFERENCE   3  (bases 1 to 4503)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..4503
                     /organism="Expression vector pColdTEV(tet)"
                     /mol_type="other DNA"
                     /db_xref="taxon:686942"
     CDS             complement(44..667)
                     /codon_start=1
                     /gene="tetR"
                     /product="tetracycline repressor"
                     /label=tetR
                     /protein_id="ACY38048.1"
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGS"
     gene            complement(44..667)
                     /gene="tetR"
                     /label=tetR
     protein_bind    692..710
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     protein_bind    722..740
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     misc_feature    762..1309
                     /label=cspA
                     /note="cspA"
     regulatory      762..828
                     /regulatory_class="promoter"
     promoter        762..828
                     /gene="cspA"
                     /label=cspA promoter
                     /note="promoter of the E. coli cold shock protein cspA gene
                     (Mitta et al., 1997)"
     misc_feature    829..849
                     /label=tet operator 1 sequence
                     /note="tet operator 1 sequence"
     protein_bind    829..847
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O1 for the tetR and tetA genes"
     5UTR           850..1002
                     /label=modified cspA 5UTR
                     /note="modified cspA 5UTR"
     5UTR           869..1002
                     /gene="cspA"
                     /label=cspA 5UTR
                     /note="5UTR of the E. coli cold shock protein cspA gene 
                     (Mitta et al., 1997)"
     misc_feature    1003..1017
                     /note="TEE; translation enhancing element"
     misc_feature    1018..1035
                     /label=His6 tag
                     /note="His6 tag"
     CDS             1018..1035
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     misc_feature    1036..1068
                     /label=TEV protease cleavage site
                     /note="TEV protease cleavage site"
     CDS             1048..1068
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     misc_feature    1069..1157
                     /note="MCS; multiple cloning site; includes NdeI, SacI, 
                     BamHI, XhoI, EcoRI, HindIII, SalI, XbaI, PstI"
     3UTR           1165..1309
                     /label=cspA 3UTR
                     /note="cspA 3UTR"
     misc_feature    complement(1472..1945)
                     /label=M13 intergenic region
                     /note="M13 intergenic region"
     rep_origin      complement(1508..1963)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     promoter        2063..2167
                     /gene="bla"
                     /label=AmpR promoter
     CDS             2168..3028
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=bla
                     /protein_id="ACY38047.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            2168..3028
                     /gene="bla"
                     /label=bla
     rep_origin      3188..3802
                     /label=ColE1 origin
                     /note="ColE1 origin"
     rep_origin      3199..3787
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"