LOCUS       Exported                6042 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pET21-vioE, complete sequence.
ACCESSION   KX461963
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6042)
  AUTHORS   Pardee K, Slomovic S, Nguyen PQ, Lee JW, Donghia N, Burrill D, 
            Ferrante T, McSorley F, Furuta Y, Lewandowski M, Boddy CN, Joshi NS,
            Collins JJ.
  TITLE     Portable, On-demand Biomolecular Manufacturing
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6042)
  AUTHORS   Pardee K, Slomovic S, Nguyen PQ, Lee JW, Donghia N, Burrill D, 
            Ferrante T, McSorley F, Furuta Y, Lewandowski M, Boddy CN, Joshi NS,
            Collins JJ.
  TITLE     Direct Submission
  JOURNAL   Submitted (29-JUN-2016) Wyss Institute, Harvard University, 3 
            Blackfan Circle, Wyss Institute Rm517-1D, Boston, MA 02115, USA
REFERENCE   3  (bases 1 to 6042)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     ##Assembly-Data-START##
            Sequencing Technology :: Sanger dideoxy sequencing 
            ##Assembly-Data-END##
FEATURES             Location/Qualifiers
     source          1..6042
                     /organism="cloning vector pET21-vioE"
                     /mol_type="other DNA"
                     /db_xref="taxon:1898576"
     rep_origin      12..467
                     /label=f1 origin
                     /note="f1 origin"
     regulatory      493..597
                     /regulatory_class="promoter"
                     /note="AmpR promoter"
     promoter        493..597
                     /gene="bla"
                     /label=AmpR promoter
     CDS             598..1458
                     /codon_start=1
                     /product="AmpR"
                     /label=AmpR
                     /protein_id="AON96401.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      1629..2217
                     /label=pBR322 origin
                     /note="pBR322 origin"
     misc_feature    2403..2545
                     /label=bom site
                     /note="bom site"
     CDS             complement(2647..2838)
                     /codon_start=1
                     /product="Rop"
                     /label=Rop
                     /protein_id="AON96402.1"
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     regulatory      complement(4729..4806)
                     /regulatory_class="promoter"
                     /note="lacI promoter"
     promoter        complement(4729..4806)
                     /gene="lacI"
                     /label=lacI promoter
     regulatory      5115..5133
                     /regulatory_class="promoter"
                     /note="T7 promoter"
     promoter        5115..5133
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    5134..5158
                     /label=lacO operator site
                     /note="lacO operator site"
     protein_bind    5134..5158
                     /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)."
     RBS             5173..5195
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             5202..5903
                     /codon_start=1
                     /product="VioE"
                     /label=VioE
                     /note="derived from Chromobacterium violaceum"
                     /protein_id="AON96403.1"
                     /translation="MGLDSSSGGGGSGGSGGGGSGENLYFQGMASDYKDDDDKHMENRE
                     PPLLPARWSSAYVSYWSPMLPDDQLTSGYCWFDYERDICRIDGLFNPWSERDTGYRLWM
                     SEVGNAASGRTWKQKVAYGRERTALGEQLCERPLDDETGPFAELFLPRDVLRRLGARHI
                     GRRVVLGREADGWRYQRPGKGPSTLYLDAASGTPLRMVTGDEASRASLRDFPNVSEAEI
                     PVFAAKREFLG"
     misc_feature    5202..5294
                     /label=N-terminal linker sequence
                     /note="N-terminal linker sequence"
     CDS             5265..5285
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     misc_feature    5295..5318
                     /label=FLAG tag
                     /note="FLAG tag"
     CDS             5295..5318
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     regulatory      5970..6017
                     /regulatory_class="terminator"
                     /note="T7 terminator"
     terminator      5970..6017
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"