LOCUS       Exported                7940 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pPBwt, complete sequence.
ACCESSION   KX573870
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7940)
  AUTHORS   Ghosh IN, Landick R.
  TITLE     OptSSeq: High-throughput sequencing readout of growth enrichment 
            defines optimal gene expression elements for homoethanologenesis
  JOURNAL   ACS Synth Biol (2016) In press
  PUBMED    27404024
REFERENCE   2  (bases 1 to 7940)
  AUTHORS   Ghosh IN, Landick RC.
  TITLE     Direct Submission
  JOURNAL   Submitted (15-JUL-2016) DOE Great Lakes Bioenergy Research Center, 
            University of Wisconsin-Madison, 1552 University Avenue, Madison, WI
            53726, United States
REFERENCE   3  (bases 1 to 7940)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..7940
                     /organism="Cloning vector pPBwt"
                     /mol_type="other DNA"
                     /db_xref="taxon:1881973"
     protein_bind    233..254
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        269..299
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     regulatory      269..298
                     /regulatory_class="promoter"
                     /note="Lac promoter"
     protein_bind    307..323
                     /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)."
     primer_bind     331..347
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             343..483
                     /codon_start=1
                     /product="LacZ alpha peptide fusion"
                     /label=LacZ alpha peptide fusion
                     /protein_id="AOH95438.1"
                     /translation="MTMITPSAQLTLTKGNKSWVPGPPSRSTVSISLISNSSSVPKNAY
                     S"
     promoter        368..386
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     416..432
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             558..2264
                     /codon_start=1
                     /product="Pdc"
                     /label=Pdc
                     /note="Z. mobilis Pyruvate Decarboxylase"
                     /protein_id="AOH95434.1"
                     /translation="MSYTVGTYLAERLVQIGLKHHFAVAGDYNLVLLDNLLLNKNMEQV
                     YCCNELNCGFSAEGYARAKGAAAAVVTYSVGALSAFDAIGGAYAENLPVILISGAPNNN
                     DHAAGHVLHHALGKTDYHYQLEMAKNITAAAEAIYTPEEAPAKIDHVIKTALREKKPVY
                     LEIACNIASMPCAAPGPASALFNDEASDEASLNAAVEETLKFIANRDKVAVLVGSKLRA
                     AGAEEAAVKFADALGGAVATMAAAKSFFPEENPHYIGTSWGEVSYPGVEKTMKEADAVI
                     ALAPVFNDYSTTGWTDIPDPKKLVLAEPRSVVVNGIRFPSVHLKDYLTRLAQKVSKKTG
                     ALDFFKSLNAGELKKAAPADPSAPLVNAEIARQVEALLTPNTTVIAETGDSWFNAQRMK
                     LPNGARVEYEMQWGHIGWSVPAAFGYAVGAPERRNILMVGDGSFQLTAQEVAQMVRLKL
                     PVIIFLINNYGYTIEVMIHDGPYNNIKNWDYAGLMEVFNGNGGYDSGAGKGLKAKTGGE
                     LAEAIKVALANTDGPTLIECFIGREDCTEELVKWGKRVAAANSRKPVNKLL"
     CDS             2324..3475
                     /codon_start=1
                     /product="AdhB"
                     /label=AdhB
                     /note="Z. mobilis Alcohol Dehydrogenase II"
                     /protein_id="AOH95435.1"
                     /translation="MASSTFYIPFVNEMGEGSLEKAIKDLNGSGFKNALIVSDAFMNKS
                     GVVKQVADLLKAQGINSAVYDGVMPNPTVTAVLEGLKILKDNNSDFVISLGGGSPHDCA
                     KAIALVATNGGEVKDYEGIDKSKKPALPLMSINTTAGTASEMTRFCIITDEVRHVKMAI
                     VDRHVTPMVSVNDPLLMVGMPKGLTAATGMDALTHAFEAYSSTAATPITDACALKAASM
                     IAKNLKTACDNGKDMPAREAMAYAQFLAGMAFNNASLGYVHAMAHQLGGYYNLPHGVCN
                     AVLLPHVLAYNASVVAGRLKDVGVAMGLDIANLGDKEGAEATIQAVRDLAASIGIPANL
                     TELGAKKEDVPLLADHALKDACALTNPRQGDQKEVEELFLSAF"
     promoter        complement(3687..3705)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3715..3731)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(4438..5112)
                     /codon_start=1
                     /product="REP"
                     /label=REP
                     /protein_id="AOH95436.1"
                     /translation="MEISMATQSREIGIQAKNKPGHWVQTERKAHEAWAGLIARKPTAA
                     MLLHHLVAQMGHQNAVVVSQKTLSKLIGRSLRTVQYAVKDLVAERWISVVKLNGPGTVS
                     AYVVNDRVAWGQPRDQLRLSVFSAAVVVDHDDQDESLLGHGDLRRIPTLYPGEQQLPTG
                     PGEEPPSQPGIPGMEPDLPALTETEEWERRGQQRLPMPDEPCFLDDGEPLEPPTRVTLP
                     RR"
     CDS             complement(4438..5100)
                     /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      5101..5870
                     /label=pBBR1 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     pBBR1 from Bordetella bronchiseptica; requires the pBBR1 
                     Rep protein for replication"
     misc_feature    5701..7098
                     /label=Mob element
                     /note="Mob element"
     CDS             complement(7153..7686)
                     /codon_start=1
                     /product="AacC1"
                     /label=AacC1
                     /note="Gentamycin-3-acetyltransferase (Gentamycin 
                     resistance)"
                     /protein_id="AOH95437.1"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPRFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        complement(7875..7903)
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"