LOCUS       Exported                7150 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Expression vector pTMH308, complete sequence.
ACCESSION   MF688773
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7150)
  AUTHORS   Hsu TM, Welner DH, Russ ZN, Cervantes B, Prathuri RL, Adams PD, 
            Dueber JE.
  TITLE     Employing a biochemical protecting group for a sustainable indigo 
            dyeing strategy
  JOURNAL   Nat. Chem. Biol. (2018) In press
  PUBMED    29309053
REFERENCE   2  (bases 1 to 7150)
  AUTHORS   Hsu TM, Welner DH, Russ ZN, Cervantes B, Prathuri RL, Adams PD, 
            Dueber JE.
  TITLE     Direct Submission
  JOURNAL   Submitted (20-AUG-2017) Bioengineering, UC Berkeley, 2151 Berkeley 
            Way Room 512D, Berkeley, CA 94704, USA
REFERENCE   3  (bases 1 to 7150)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..7150
                     /organism="Expression vector pTMH308"
                     /mol_type="other DNA"
                     /db_xref="taxon:2070485"
     misc_feature    168..310
                     /label=bom
                     /note="basis of mobility region from pBR322"
     CDS             complement(412..603)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
     CDS             complement(1915..2997)
                     /codon_start=1
                     /product="LacI lac repressor"
                     /label=LacI lac repressor
                     /protein_id="AUO17149.1"
                     /translation="MKPVTLYDVAEYAGVSYQTVSRVVNQASHVSAKTREKVEAAMAEL
                     NYIPNRVAQQLAGKQSLLIGVATSSLALHAPSQIVAAIKSRADQLGASVVVSMVERSGV
                     EACKAAVHNLLAQRVSGLIINYPLDDQDAIAVEAACTNVPALFLDVSDQTPINSIIFSH
                     EDGTRLGVEHLVALGHQQIALLAGPLSSVSARLRLAGWHKYLTRNQIQPIAEREGDWSA
                     MSGFQQTMQMLNEGIVPTAMLVANDQMALGAMRAITESGLRVGADISVVGYDDTEDSSC
                     YIPPLTTIKQDFRLLGQTSVDRLLQLSQGQAVKGNQLLPVSLVKRKTTLAPNTQTASPR
                     ALADSLMQLARQVSRLESGQ"
     promoter        complement(2998..3075)
                     /gene="lacI"
                     /label=lacI promoter
     regulatory      3384..3403
                     /regulatory_class="promoter"
                     /note="T7"
     promoter        3384..3402
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     protein_bind    3403..3427
                     /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)."
     protein_bind    3403..3425
                     /label=LacI binding site
                     /bound_moiety="LacI"
                     /note="LacO lac operator"
     RBS             3442..3464
                     /note="efficient ribosome binding site from bacteriophage 
                     T7 gene 10 (Olins and Rangwala, 1989)"
     CDS             3471..4970
                     /codon_start=1
                     /product="PtUGT72B30 indoxyl glucosyltransferase"
                     /label=PtUGT72B30 indoxyl glucosyltransferase
                     /note="from Polygonum tinctorium with N-terminal 6xHis tag"
                     /protein_id="AUO17150.1"
                     /translation="MGHHHHHHDYDIPTTENLYFQGSESPAAPPTTAPPPHVIIMPSAG
                     MGHLIPLAEFAKRLLPRFTFTFAVPTSGPPSSSQRDFLSSLPASIDTSFLPEVDLSDAP
                     SDAQIETLMSLMVVRSLPSLRDLIASYSASGRRVAALVVDLFATDAIDVALELGIRPFI
                     FFPSTAMTLSFFLHLEKLDETVSCEFAELSDPVQIPGCIPVHGKDLIDPVQDRKNDAYK
                     WLLHHSKRYKLAEGVIVNSFEGLEAGPIRQLLHPEPGKPRVYPVGPLIQAGSCEKGAAA
                     RPECLKWLDQQPRGSVLFVNFGSGGVLSTEQQNELAGVLAHSQQRFLWVVRPPNDGIAN
                     ATYFSVDGEIDPLKLLPEGFLEQTAGRGLVLPMWAPQIDVLSHESTGGFLTHCGWNSTL
                     ESVFHGVPLITWPLYAEQKMNAVMLTEGLRVGLRPSVGKDGIIRGDEIARVIGELMEGE
                     EGKRIRSKMQELKRAASAVLSKDGSSTRALEEVAKIWESKV"
     CDS             3477..3494
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             3516..3536
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQG"
     misc_feature    3540..4967
                     /note="PtUGT72B30 indoxyl glucosyltransferase from 
                     Polygonum tinctorium"
     regulatory      5045..5092
                     /regulatory_class="terminator"
                     /note="T7 terminator"
     terminator      5045..5092
                     /label=T7 terminator
                     /note="transcription terminator for bacteriophage T7 RNA 
                     polymerase"
     promoter        complement(5267..5295)
                     /label=tet promoter
                     /note="E. coli promoter for tetracycline efflux protein 
                     gene"
     regulatory      5382..5512
                     /regulatory_class="promoter"
     promoter        5408..5512
                     /gene="bla"
                     /label=AmpR promoter
     CDS             5513..6373
                     /codon_start=1
                     /product="ampicillin resistance protein"
                     /label=ampicillin resistance protein
                     /protein_id="AUO17151.1"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRVDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPAAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     regulatory      6371..6467
                     /regulatory_class="terminator"
     rep_origin      6468..7150
                     /label=ColE1 origin
                     /note="ColE1 origin"
     rep_origin      6544..7132
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"