LOCUS       Exported               14206 bp ds-DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pKL13, complete sequence.
ACCESSION   KU746975
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 14206)
  AUTHORS   Lam KN.
  TITLE     Developing a Bacteroides system for function-based screening of DNA 
            from the human gut microbiome
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 14206)
  AUTHORS   Lam KN.
  TITLE     Direct Submission
  JOURNAL   Submitted (22-FEB-2016) Biology, University of Waterloo, 200 
            University Avenue West, Waterloo, Canada
REFERENCE   3  (bases 1 to 14206)
  AUTHORS   .
  TITLE     Direct Submission
FEATURES             Location/Qualifiers
     source          1..14206
                     /organism="Cloning vector pKL13"
                     /mol_type="other DNA"
                     /country="Canada"
                     /db_xref="taxon:1898397"
     primer_bind     288..304
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      311..329
                     /regulatory_class="promoter"
     promoter        311..329
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     misc_feature    332..337
                     /label=EcoRI site
                     /note="EcoRI site"
     CDS             complement(931..1620)
                     /codon_start=1
                     /gene="repA"
                     /product="replication protein"
                     /label=repA
                     /protein_id="AOP12487.1"
                     /translation="MILNRKIIFPLILFLWKIIVYFVENNISQKHIKMENKKAVKLTDF
                     QKNEENPFMKQAIEGIENHVVKKYKSNSGGDKRAVVALADTETGEVFKTSFIRQIEVDE
                     EQFTKLYLSNFAAFFDLSQAAIRVFGYFMTCMKPKNDLIIFNRKKCLEYTKYKTDKAVY
                     KGLAELVKAEIIARGPADNLWFINPLIVFNGDRVTFAKTYVRKKTLAAQKKEEAEKRQL
                     SLGFDEQ"
     gene            complement(931..1620)
                     /gene="repA"
                     /label=repA
     CDS             3183..3983
                     /codon_start=1
                     /gene="ermF"
                     /product="erythromycin resistance"
                     /label=ermF
                     /protein_id="AOP12484.1"
                     /translation="MTKKKLPVRFTGQHFTIDKVLIKDAIRQANISNQDTVLDIGAGKG
                     FLTVHLLKIANNVVAIENDTALVEHLRKLFSDARNVQVVGCDFRNFAVPKFPFKVVSNI
                     PYGITSDIFKILMFESLGNFLGGSIVLQLEPTQKLFSRKLYNPYTVFYHTFFDLKLVYE
                     VGPESFLPPPTVKSALLNIKRKHLFFDFKFKAKYLAFISCLLEKPDLSVKTALKSIFRK
                     SQVRSISEKFGLNLNAQIVCLSPSQWLNCFLEMLEVVPEKFHPS"
     gene            3183..3983
                     /gene="ermF"
                     /label=ermF
     misc_feature    4133..4138
                     /label=EcoRI site
                     /note="EcoRI site"
     misc_feature    4167..4174
                     /label=SgsI site
                     /note="SgsI site"
     misc_feature    4183..4190
                     /label=PacI site
                     /note="PacI site"
     regulatory      complement(4191..4279)
                     /regulatory_class="terminator"
                     /note="ilvGEDA transcriptional terminator"
     primer_bind     4280..4310
                     /label=KL-JC103 sequencing primer
                     /note="KL-JC103 sequencing primer"
     misc_feature    complement(4311..4321)
                     /label=3-frame translational stop 2
                     /note="3-frame translational stop 2"
     misc_feature    4322..4327
                     /label=NheI site
                     /note="NheI site"
     misc_feature    4328..4333
                     /label=Eco72I site
                     /note="Eco72I site"
     misc_feature    4334..4340
                     /label=Eco81I site
                     /note="Eco81I site"
     regulatory      complement(4341..4407)
                     /regulatory_class="promoter"
                     /note="tac promoter operator"
     protein_bind    4349..4365
                     /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(4373..4401)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     misc_feature    4408..4414
                     /label=Eco81I site
                     /note="Eco81I site"
     misc_feature    4415..4420
                     /label=BstBI site
                     /note="BstBI site"
     CDS             4607..5407
                     /codon_start=1
                     /gene="aac(3)IV"
                     /product="aminoglycoside 3-N-acetyltransferase type IV     
                               "
                     /label=ApmR
                     /note="confers resistance to apramycin (Paget and Davies, 
                     1996)"
                     /translation="MSSAVECNVVQYEWRKAELIGQLLNLGVTPGGVLLVHSSFRSVRP
                     LEDGPLGLIEALRAALGPGGTLVMPSWSGLDDEPFDPATSPVTPDLGVVSDTFWRLPNV
                     KRSAHPFAFAAAGPQAEQIISDPLPLPPHSPASPVARVHELDGQVLLLGVGHDANTTLH
                     LAELMAKVPYGVPRHCTILQDGKLVRVDYLENDHCCERFALADRWLKEKSLQKEGPVGH
                     AFARLIRSRDIVATALGQLGRDPLIFLHPPEAGCEECDAARQSIG"
     CDS             4634..5410
                     /codon_start=1
                     /product="gentamicin resistance protein"
                     /label=gentamicin resistance protein
                     /protein_id="AOP12485.1"
                     /translation="MQYEWRKAELIGQLLNLGVTPGGVLLVHSSFRSVRPLEDGPLGLI
                     EALRAALGPGGTLVMPSWSGLDDEPFDPATSPVTPDLGVVSDTFWRLPNVKRSAHPFAF
                     AAAGPQAEQIISDPLPLPPHSPASPVARVHELDGQVLLLGVGHDANTTLHLAELMAKVP
                     YGVPRHCTILQDGKLVRVDYLENDHCCERFALADRWLKEKSLQKEGPVGHAFARLIRSR
                     DIVATALGQLGRDPLIFLHPPEAGCEECDAARQSIG"
     misc_feature    5476..5481
                     /label=BstBI site
                     /note="BstBI site"
     misc_feature    5482..5487
                     /label=Eco72I site
                     /note="Eco72I site"
     misc_feature    5488..5493
                     /label=NsiI site
                     /note="NsiI site"
     misc_feature    5494..5504
                     /label=3-frame translational stop 1
                     /note="3-frame translational stop 1"
     primer_bind     complement(5505..5535)
                     /label=KL-JC102 sequencing primer
                     /note="KL-JC102 sequencing primer"
     primer_bind     complement(5506..5522)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     regulatory      5536..5617
                     /regulatory_class="terminator"
                     /note="rnpB T1 transcriptional terminator"
     misc_feature    5618..5624
                     /label=CpoI site
                     /note="CpoI site"
     misc_feature    5633..5640
                     /label=SfaAI site
                     /note="SfaAI site"
     misc_feature    5684..6461
                     /label=RK2 oriT region
                     /note="RK2 oriT region"
     oriT            5822..5931
                     /note="incP origin of transfer"
     CDS             5964..6335
                     /codon_start=1
                     /gene="traJ"
                     /product="oriT-recognizing protein"
                     /label=traJ
                     /translation="MADETKPTRKGSPPIKVYCLPDERRAIEEKAAAAGMSLSAYLLAV
                     GQGYKITGVVDYEHVRELARINGDLGRLGGLLKLWLTDDPRTARFGDATILALLAKIEE
                     KQDELGKVMMGVVRPRAEP"
     primer_bind     complement(6510..6526)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    6534..6550
                     /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(6558..6588)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     regulatory      complement(6559..6588)
                     /regulatory_class="promoter"
                     /note="lac promoter"
     protein_bind    6603..6624
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     CDS             complement(6844..7503)
                     /codon_start=1
                     /gene="CAT"
                     /product="chloramphenicol resistance protein"
                     /label=CAT
                     /protein_id="AOP12488.1"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     gene            complement(6844..7503)
                     /gene="CAT"
                     /label=CAT
     promoter        complement(7504..7606)
                     /label=cat promoter
                     /note="promoter of the E. coli cat gene encoding 
                     chloramphenicol acetyltransferase"
     CDS             7722..8069
                     /codon_start=1
                     /gene="redF"
                     /product="RedF"
                     /label=redF
                     /protein_id="AOP12489.1"
                     /translation="MERRNRRTGRTEKARIWEVTDRTVRTWIGEAVAAAAADGVTFSVP
                     VTPHTFRHSYAMHMLYAGIPLKVLQSLMGHKSISSTEVYTKVFALDVAARHRVQFAMPE
                     SDAVAMLKQLS"
     gene            7722..8069
                     /gene="redF"
                     /label=redF
     rep_origin      8464..9078
                     /label=RK2 oriV
                     /note="RK2 oriV"
     rep_origin      9154..9373
                     /label=ori2
                     /note="secondary origin of replication for the bacterial F 
                     plasmid; also known as oriS"
     rep_origin      9154..9370
                     /label=F plasmid origin
                     /note="F plasmid origin"
     CDS             9464..10219
                     /codon_start=1
                     /gene="repE"
                     /product="replication initiation protein"
                     /label=repE
                     /protein_id="AOP12486.1"
                     /translation="MAETAVINHKKRKNSPRIVQSNDLTEAAYSLSRDQKRMLYLFVDQ
                     IRKSDGTLQEHDGICEIHVAKYAEIFGLTSAEASKDIRQALKSFAGKEVVFYRPEEDAG
                     DEKGYESFPWFIKRAHSPSRGLYSVHINPYLIPFFIGLQNRFTQFRLSETKEITNPYAM
                     RLYESLCQYRKPDGSGIVSLKIDWIIERYQLPQSYQRMPDFRRRFLQVCVNEINSRTPM
                     RLSYIEKKKGRQTTHIVFSFRDITSMTTG"
     gene            9464..10219
                     /gene="repE"
                     /label=repE
     misc_feature    10222..10472
                     /gene="incC"
                     /label=incC
                     /note="incompatibility region of the bacterial F plasmid"
     CDS             10798..11973
                     /codon_start=1
                     /gene="parA"
                     /product="partitioning protein"
                     /label=parA
                     /protein_id="AOP12482.1"
                     /translation="MFRMKLMETLNQCINAGHEMTKAIAIAQFNDDSPEARKITRRWRI
                     GEAADLVGVSSQAIRDAEKAGRLPHPDMEIRGRVEQRVGYTIEQINHMRDVFGTRLRRA
                     EDVFPPVIGVAAHKGGVYKTSVSVHLAQDLALKGLRVLLVEGNDPQGTASMYHGWVPDL
                     HIHAEDTLLPFYLGEKDDVTYAIKPTCWPGLDIIPSCLALHRIETELMGKFDEGKLPTD
                     PHLMLRLAIETVAHDYDVIVIDSAPNLGIGTINVVCAADVLIVPTPAELFDYTSALQFF
                     DMLRDLLKNVDLKGFEPDVRILLTKYSNSNGSQSPWMEEQIRDAWGSMVLKNVVRETDE
                     VGKGQIRMRTVFEQAIDQRSSTGAWRNALSIWEPVCNEIFDRLIKPRWEIR"
     gene            10798..11973
                     /gene="parA"
                     /label=parA
     CDS             11973..12944
                     /codon_start=1
                     /gene="parB"
                     /product="partitioning protein"
                     /label=parB
                     /protein_id="AOP12483.1"
                     /translation="MKRAPVIPKHTLNTQPVEDTSLSTPAAPMVDSLIARVGVMARGNA
                     ITLPVCGRDVKFTLEVLRGDSVEKTSRVWSGNERDQELLTEDALDDLIPSFLLTGQQTP
                     AFGRRVSGVIEIADGSRRRKAAALTESDYRVLVGELDDEQMAALSRLGNDYRPTSAYER
                     GQRYASRLQNEFAGNISALADAENISRKIITRCINTAKLPKSVVALFSHPGELSARSGD
                     ALQKAFTDKEELLKQQASNLHEQKKAGVIFEAEEVITLLTSVLKTSSASRTSLSSRHQF
                     APGATVLYKGDKMVLNLDRSRVPTECIEKIEAILKELEKPAP"
     gene            11973..12944
                     /gene="parB"
                     /label=parB
     misc_feature    13006..13555
                     /label=parC region
                     /note="parC region"
     misc_feature    13017..13490
                     /gene="sopC"
                     /label=sopC
                     /note="centromere-like partitioning region of the bacterial
                     F plasmid"
     misc_feature    13750..14148
                     /label=cos
                     /note="lambda cos site; allows packaging into phage lambda 
                     particles"
     misc_feature    13953..13965
                     /label=cos site
                     /note="cos site"
     misc_recomb     14166..14199
                     /label=loxP
                     /note="loxP"
     protein_bind    14166..14199
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (GCATACAT)."