LOCUS       40924_36298        5953 bp DNA     circular SYN 18-DEC-2018
DEFINITION  Cloning vector pR6KT2, complete sequence.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5953)
  AUTHORS   Kvitko BH, Bruckbauer S, Prucha J, McMillan I, Breland EJ, Lehman S,
            Mladinich K, Choi KH, Karkhoff-Schweizer R, Schweizer HP.
  TITLE     A simple method for construction of pir+ Enterobacterial hosts for 
            maintenance of R6K replicon plasmids
  JOURNAL   BMC Res Notes 5, 157 (2012)
  PUBMED    22433797
REFERENCE   2  (bases 1 to 5953)
  AUTHORS   Bruckbauer ST.
  TITLE     Direct Submission
  JOURNAL   Submitted (25-JUN-2012) Microbiology Immunology and Pathology, 
            Colorado State University, IDRC at Foothills Campus, Campus Delivery
            0922, Fort Collins, CO 80523-0922, USA
REFERENCE   3  (bases 1 to 5953)
  TITLE     Direct Submission
REFERENCE   4  (bases 1 to 5953)
  AUTHORS   .
  TITLE     Direct Submission
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "BMC Res 
            Notes 5, 157 (2012)"
COMMENT     SGRef: number: 2; type: "Journal Article"; journalName: "Submitted 
            (25-JUN-2012) Microbiology Immunology and Pathology, Colorado State 
            University, IDRC at Foothills Campus, Campus Delivery 0922, Fort 
            Collins, CO 80523-0922, USA"
COMMENT     SGRef: number: 3; type: "Journal Article"
FEATURES             Location/Qualifiers
     source          1..5953
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     269..285
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     rep_origin      complement(441..825)
                     /direction=LEFT
                     /label=R6K gamma ori
                     /note="gamma replication origin from E. coli plasmid R6K;
                     requires the R6K initiator protein pi for replication"
     oriT            complement(848..956)
                     /direction=LEFT
                     /label=oriT
                     /note="incP origin of transfer"
     promoter        1306..1334
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     CDS             1523..2053
                     /codon_start=1
                     /label=GmR
                     /note="gentamycin acetyltransferase"
                     /translation="MLRSSNDVTQQGSRPKTKLGGSSMGIIRTCRLGPDQVKSMRAALD
                     LFGREFGDVATYSQHQPDSDYLGNLLRSKTFIALAAFDQEAVVGALAAYVLPRFEQPRS
                     EIYIYDLAVSGEHRRQGIATALINLLKHEANALGAYVIYVQADYGDDPAVALYTKLGIR
                     EEVMHFDIDPSTAT"
     promoter        2152..2180
                     /gene="intI1 (promoter lies within the coding sequence)"
                     /label=Pc promoter
                     /note="class 1 integron promoter"
     CDS             2506..4314
                     /codon_start=1
                     /gene="uidA"
                     /product="beta-D-glucuronidase"
                     /label=uidA
                     /note="Breaks down X-Gluc producing a blue phenotype"
                     /protein_id="AFR24688.1"
                     /translation="MLRPVETPTREIKKLDGLWAFSLDRENCGIDQRWWESALQESRAI
                     AVPGSFNDQFADADIRNYAGNVWYQREVFIPKGWAGQRIVLRFDAVTHYGKVWVNNQEV
                     MEHQGGYTPFEADVTPYVIAGKSVRITVCVNNELNWQTIPPGMVITDENGKKKQSYFHD
                     FFNYAGIHRSVMLYTTPNTWVDDITVVTHVAQDCNHASVDWQVVANGDVSVELRDADQQ
                     VVATGQGTSGTLQVVNPHLWQPGEGYLYELCVTAKSQTECDIYPLRVGIRSVAVKGEQF
                     LINHKPFYFTGFGRHEDADLRGKGFDNVLMVHDHALMDWIGANSYRTSHYPYAEEMLDW
                     ADEHGIVVIDETAAVGFNLSLGIGFEAGNKPKELYSEEAVNGETQQAHLQAIKELIARD
                     KNHPSVVMWSIANEPDTRPQVHGNISPLAEATRKLDPTRPITCVNVMFCDAHTDTISDL
                     FDVLCLNRYYGWYVQSGDLETAEKVLEKELLAWQEKLHQPIIITEYGVDTLAGLHSMYT
                     DMWSEEYQCAWLDMYHRVFDRVSAVVGEQVWNFADFATSQGILRVGGNKKGIFTRDRKP
                     KSAAFLLQKRWTGMNFGEKPQQGGKQ"
     gene            2506..4314
                     /gene="uidA"
                     /label=uidA
     terminator      4370..4413
                     /label=rrnB T1 terminator
                     /note="transcription terminator T1 from the E. coli rrnB
                     gene"
     CDS             complement(4430..5782)
                     /codon_start=1
                     /label=SacB
                     /note="secreted levansucrase that renders bacterial growth 
                     sensitive to sucrose"
                     /translation="GATQAFAKETNQKPYKETYGISHITRHDMLQIPEQQKNEKYQVSE
                     FDSSTIKNISSAKGLDVWDSWPLQNADGTVANYHGYHIVFALAGDPKNADDTSIYMFYQ
                     KVGETSIDSWKNAGRVFKDSDKFDANDSILKDQTQEWSGSATFTSDGKIRLFYTDFSGK
                     HYGKQTLTTAQVNVSASDSSLNINGVEDYKSIFDGDGKTYQNVQQFIDEGNYSSGDNHT
                     LRDPHYVEDKGHKYLVFEANTGTEDGYQGEESLFNKAYYGKSTSFFRQESQKLLQSDKK
                     RTAELANGALGMIELNDDYTLKKVMKPLIASNTVTDEIERANVFKMNGKWYLFTDSRGS
                     KMTIDGITSNDIYMLGYVSNSLTGPYKPLNKTGLVLKMDLDPNDVTFTYSHFAVPQAKG
                     NNVVITSYMTNRGFYADKQSTFAPSFLLNIKGKKTSVVKDSILEQGQLTVNK"
ORIGIN
        1 cgatcctcta gctagagtcg atcttcgcca gcagggcgag gatcgtggca tcaccgaacc
       61 gcgccgtgcg cgggtcgtcg gtgagccaga gtttcagcag gccgcccagg cggcccaggt
      121 cgccattgat gcgggccagc tcgcggacgc aactgttggg aagggcgatc ggtgcgggcc
      181 tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta
      241 acgccagggt tttcccagtc acgacgttgt aaaacgacgg ccagtgccaa gctcattacc
      301 ctgttatccc tacccctcgg atccactagt aacggccgcc agtgtgctgg aattcggctt
      361 taatcccggg aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc
      421 taatcctgtt accagccggt tgtcagccgt taagtgttcc tgtgtcactc aaaattgctt
      481 tgagaggctc taagggcttc tcagtgcgtt acatccctgg cttgttgtcc acaaccgtta
      541 aaccttaaaa gctttaaaag ccttatatat tctttttttt cttataaaac ttaaaacctt
      601 agaggctatt taagttgctg atttatatta attttattgt tcaaacatga gagcttagta
      661 cgtgaaacat gagagcttag tacgttagcc atgagagctt agtacgttag ccatgagggt
      721 ttagttcgtt aaacatgaga gcttagtacg ttaaacttga gagcttagta cgtgaaacat
      781 gagagcttag tacgtactat caacaggttg aactgcccat gttctttcct gcgttatcag
      841 agcttatcgg ccagcctcgc agagcaggat tcccgttgag caccgccagg tgcgaataag
      901 ggacagtgaa gaaggaacac ccgctcgcgg gtgggcctac ttcacctatc ctgcccggct
      961 gacgccgttg gatacaccaa ggaaagtcta cacgaaccct ttggcaaaat cctgtatatc
     1021 gtgcgaaaaa ggatggatat accgaaaaaa tcgctataat gaccccgaag cagggttatg
     1081 cagcggaaag tataccttaa ggaatcccct gataccgctc gccgcagccg aacgaccgag
     1141 cgcagcgggt cagtgagcga ggaagcggaa gcccgggaat aagccgaatt ctgcagatat
     1201 ccatcacact ggcggccgct cgagcatgca tctagagaga ataggaactt cggaatagga
     1261 acttcaagat cccctgattc cctttgtcaa cagcaatgga tcgaattgac ataagcctgt
     1321 tcggttcgta aactgtaatg caagtagcgt atgcgctcac gcaactggtc cagaaccttg
     1381 accgaacgca gcggtggtaa cggcgcagtg gcggttttca tggcttgtta tgactgtttt
     1441 tttgtacagt ctatgcctcg ggcatccaag cagcaagcgc gttacgccgt gggtcgatgt
     1501 ttgatgttat ggagcagcaa cgatgttacg cagcagcaac gatgttacgc agcagggcag
     1561 tcgccctaaa acaaagttag gtggctcaag tatgggcatc attcgcacat gtaggctcgg
     1621 ccctgaccaa gtcaaatcca tgcgggctgc tcttgatctt ttcggtcgtg agttcggaga
     1681 cgtagccacc tactcccaac atcagccgga ctccgattac ctcgggaact tgctccgtag
     1741 taagacattc atcgcgcttg ctgccttcga ccaagaagcg gttgttggcg ctctcgcggc
     1801 ttacgttctg cccaggtttg agcagccgcg tagtgagatc tatatctatg atctcgcagt
     1861 ctccggcgag caccggaggc agggcattgc caccgcgctc atcaatctcc tcaagcatga
     1921 ggccaacgcg cttggtgctt atgtgatcta cgtgcaagca gattacggtg acgatcccgc
     1981 agtggctctc tatacaaagt tgggcatacg ggaagaagtg atgcactttg atatcgaccc
     2041 aagtaccgcc acctaacaat tcgttcaagc cgagatcggc ttcccggccg cggagttgtt
     2101 cggtaaattg tcacaacgcc gcggccaatt cgatagtgaa cacgaaccca gttgacataa
     2161 gcctgttcgg ttcgtaaact gtaatgcaag tagcgtatgc gctcacgcaa ctggtccaga
     2221 accttgaccg aacgcagcgg tggtaacggc gcagtggcgg ttttcatggc ttgttatgac
     2281 tgtttttttg tacagtctat gcctcgggca tccaagcagc aagcgcgtta cgccgtgggt
     2341 cgatgtttga tgttatggag cagcaacgat gttacgcagc agggcagtcg ccctaaaaca
     2401 aagttaggca gccgttgtgc tggtgctttc tgatagttgt tgtggggtag gcagtcagag
     2461 ctcgatttgc ttgtcgccat aatagattca caagaaggat tcgacatgtt acgtcctgta
     2521 gaaaccccaa cccgtgaaat caaaaaactc gacggcctgt gggcattcag tctggatcgc
     2581 gaaaactgtg gaattgatca gcgttggtgg gaaagcgcgt tacaagaaag ccgggcaatt
     2641 gctgtgccag gcagttttaa cgatcagttc gccgatgcag atattcgtaa ttatgcgggc
     2701 aacgtctggt atcagcgcga agtctttata ccgaaaggtt gggcaggcca gcgtatcgtg
     2761 ctgcgtttcg atgcggtcac tcattacggc aaagtgtggg tcaataatca ggaagtgatg
     2821 gagcatcagg gcggctatac gccatttgaa gccgatgtca cgccgtatgt tattgccggg
     2881 aaaagtgtac gtatcaccgt ttgtgtgaac aacgaactga actggcagac tatcccgccg
     2941 ggaatggtga ttaccgacga aaacggcaag aaaaagcagt cttacttcca tgatttcttt
     3001 aactatgccg ggatccatcg cagcgtaatg ctctacacca cgccgaacac ctgggtggac
     3061 gatatcaccg tggtgacgca tgtcgcgcaa gactgtaacc acgcgtctgt tgactggcag
     3121 gtggtggcca atggtgatgt cagcgttgaa ctgcgtgatg cggatcaaca ggtggttgca
     3181 actggacaag gcactagcgg gactttgcaa gtggtgaatc cgcacctctg gcaaccgggt
     3241 gaaggttatc tctatgaact gtgcgtcaca gccaaaagcc agacagagtg tgatatctac
     3301 ccgcttcgcg tcggcatccg gtcagtggca gtgaagggcg aacagttcct gattaaccac
     3361 aaaccgttct actttactgg ctttggtcgt catgaagatg cggacttacg tggcaaagga
     3421 ttcgataacg tgctgatggt gcacgaccac gcattaatgg actggattgg ggccaactcc
     3481 taccgtacct cgcattaccc ttacgctgaa gagatgctcg actgggcaga tgaacatggc
     3541 atcgtggtga ttgatgaaac tgctgctgtc ggctttaacc tctctttagg cattggtttc
     3601 gaagcgggca acaagccgaa agaactgtac agcgaagagg cagtcaacgg ggaaactcag
     3661 caagcgcact tacaggcgat taaagagctg atagcgcgtg acaaaaacca cccaagcgtg
     3721 gtgatgtgga gtattgccaa cgaaccggat acccgtccgc aagtgcacgg gaatatttcg
     3781 ccactggcgg aagcaacgcg taaactcgac ccgacgcgtc cgatcacctg cgtcaatgta
     3841 atgttctgcg acgctcacac cgataccatc agcgatctct ttgatgtgct gtgcctgaac
     3901 cgttattacg gatggtatgt ccaaagcggc gatttggaaa cggcagagaa ggtactggaa
     3961 aaagaacttc tggcctggca ggagaaactg catcagccga ttatcatcac cgaatacggc
     4021 gtggatacgt tagccgggct gcactcaatg tacaccgaca tgtggagtga agagtatcag
     4081 tgtgcatggc tggatatgta tcaccgcgtc tttgatcgcg tcagcgccgt cgtcggtgaa
     4141 caggtatgga atttcgccga ttttgcgacc tcgcaaggca tattgcgcgt tggcggtaac
     4201 aagaaaggga tcttcactcg cgaccgcaaa ccgaagtcgg cggcttttct gctgcaaaaa
     4261 cgctggactg gcatgaactt cggtgaaaaa ccgcagcagg gaggcaaaca atgaatcaac
     4321 aactctcctg gcgcaccatc gtcggctaca gcctcggtga cgtctcagat taaaacgaaa
     4381 ggcccagtct ttcgactgag cctttcgttt tatttgaaag ctactattat ttgttaactg
     4441 ttaattgtcc ttgttcaagg atgctgtctt tgacaacaga tgttttcttg cctttgatgt
     4501 tcagcaggaa gctcggcgca aacgttgatt gtttgtctgc gtagaatcct ctgtttgtca
     4561 tatagcttgt aatcacgaca ttgtttcctt tcgcttgagg tacagcgaag tgtgagtaag
     4621 taaaggttac atcgttagga tcaagatcca tttttaacac aaggccagtt ttgttcagcg
     4681 gcttgtatgg gccagttaaa gaattagaaa cataaccaag catgtaaata tcgttagacg
     4741 taatgccgtc aatcgtcatt tttgatccgc gggagtcagt gaacaggtac catttgccgt
     4801 tcattttaaa gacgttcgcg cgttcaattt catctgttac tgtgttagat gcaatcagcg
     4861 gtttcatcac ttttttcagt gtgtaatcat cgtttagctc aatcataccg agagcgccgt
     4921 ttgctaactc agccgtgcgt tttttatcgc tttgcagaag tttttgactt tcttgacgga
     4981 agaatgatgt gcttttgcca tagtatgctt tgttaaataa agattcttcg ccttggtagc
     5041 catcttcagt tccagtgttt gcttcaaata ctaagtattt gtggccttta tcttctacgt
     5101 agtgaggatc tctcagcgta tggttgtcgc ctgagctgta gttgccttca tcgatgaact
     5161 gctgtacatt ttgatacgtt tttccgtcac cgtcaaagat tgatttataa tcctctacac
     5221 cgttgatgtt caaagagctg tctgatgctg atacgttaac ttgtgcagtt gtcagtgttt
     5281 gtttgccgta atgtttaccg gagaaatcag tgtagaataa acggattttt ccgtcagatg
     5341 taaatgtggc tgaacctgac cattcttgtg tttggtcttt taggatagaa tcatttgcat
     5401 cgaatttgtc gctgtcttta aagacgcggc cagcgttttt ccagctgtca atagaagttt
     5461 cgccgacttt ttgatagaac atgtaaatcg atgtgtcatc cgcattttta ggatctccgg
     5521 ctaatgcaaa gacgatgtgg tagccgtgat agtttgcgac agtgccgtca gcgttttgta
     5581 atggccagct gtcccaaacg tccaggcctt ttgcagaaga gatattttta attgtggacg
     5641 aatcaaattc agaaacttga tatttttcat ttttttgctg ttcagggatt tgcagcatat
     5701 catggcgtgt aatatgggaa atgccgtatg tttccttata tggcttttgg ttcgtttctt
     5761 tcgcaaacgc ttgagttgcg ccgtcgctct gcgcgtgcgc cgacatgcaa gcggctgcgg
     5821 tgagcgcggc ggcgctcagg gccaaacgtt tgtgtctacg ggaaacggct tgtttggagt
     5881 acgtcatcgt ttcctcctgt gcagcgagtg ctaattatat ggacgggttt tgacaaattc
     5941 aagaagtgac tag
//
