LOCUS       Exported                5236 bp ds-DNA     circular SYN 17-DEC-2018
DEFINITION  Expression vector LIC-pIVEX-LC2, complete sequence.
ACCESSION   JF327845
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5236)
  AUTHORS   Dortay H, Akula UM, Westphal C, Sittig M, Mueller-Roeber B.
  TITLE     High-Throughput Protein Expression Using a Combination of 
            Ligation-Independent Cloning (LIC) and Infrared Fluorescent Protein 
            (IFP) Detection
  JOURNAL   PLoS ONE 6 (4), E18900 (2011)
  PUBMED    21541323
REFERENCE   2  (bases 1 to 5236)
  AUTHORS   Dortay H, Mueller-Roeber B.
  TITLE     Direct Submission
  JOURNAL   Submitted (12-FEB-2011) Molecular Biology, University of Potsdam, 
            Germany, Karl-Liebknecht-Str. 24-25, Haus 20, Potsdam, Brandenburg 
            14476, Germany
REFERENCE   3  (bases 1 to 5236)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..5236
                     /organism="Expression vector LIC-pIVEX-LC2"
                     /mol_type="other DNA"
                     /db_xref="taxon:1029638"
     primer_bind     381..397
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     promoter        620..638
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      620..636
                     /regulatory_class="promoter"
                     /note="T7 promoter"
     RBS             670..692
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     regulatory      686..691
                     /regulatory_class="ribosome_binding_site"
     misc_feature    699..701
                     /label=start codon
                     /note="start codon"
     misc_feature    711..718
                     /label=PmeI recognition site
                     /note="PmeI recognition site"
     misc_feature    719..1385
                     /label=LIC stuffer fragment
                     /note="LIC stuffer fragment"
     misc_feature    1386..1393
                     /label=PmeI recognition site
                     /note="PmeI recognition site"
     misc_feature    1399..1419
                     /label=TEV recognition site
                     /note="TEV recognition site"
     CDS             1399..1419
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     misc_feature    1429..2412
                     /label=infrared fluorescent protein
                     /note="infrared fluorescent protein"
     CDS             1429..2412
                     /codon_start=1
                     /product="bacteriophytochrome-based monomeric infrared 
                     fluorescent protein (Shu et al., 2009)"
                     /label=IFP1.4
                     /translation="MARDPLPFFPPLYLGGPEITTENCEREPIHIPGSIQPHGALLTAD
                     GHSGEVLQVSLNAATFLGQEPTVLRGQTLAALLPEQWPALQAALPPGCPDALQYRATLD
                     WPAAGHLSLTVHRVAELLILEFEPTEAWDSIGPHALRNAMFALESAPNLRALAEVATQT
                     VRELTGFDRVMLYKFAPDATGEMIAEARREGMQAFLGHRFPASHTPAQARALYTRHLLR
                     LTADTRAAAVPLDPVLNPQTNAPTPLGGAVLRATSPMHMQYLRNMGVGSSLSVSVVVGG
                     QLWGLIVCHHQTPYVLPPDLRTTLEELGRKLSGQVQRKEAGMDELYK"
     misc_feature    2428..2448
                     /label=6xHis tag
                     /note="6xHis tag"
     CDS             2428..2445
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     misc_feature    2446..2451
                     /label=double stop codon (TAA TAA)
                     /note="double stop codon (TAA TAA)"
     terminator      2553..2600
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     regulatory      2560..2597
                     /regulatory_class="terminator"
                     /note="T7 terminator"
     promoter        complement(2966..2994)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     primer_bind     complement(3015..3031)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3039..3055
                     /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)."
     promoter        complement(3063..3093)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3108..3129
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      3358..4031
                     /label=pBR322 origin
                     /note="pBR322 origin"
     rep_origin      complement(3417..4005)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(4176..5036)
                     /codon_start=1
                     /gene="ampR"
                     /product="beta-lactamase"
                     /label=ampR
                     /protein_id="AEE81069.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     gene            complement(4176..5036)
                     /gene="ampR"
                     /label=ampR
     promoter        complement(5037..5141)
                     /gene="bla"
                     /label=AmpR promoter