LOCUS       sequence-234368-        8164 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Broad host range plasmid. Theophylline inducible expression of YFP 
            in various cyanobacteria..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8164)
  AUTHORS   Ma AT, Schmidt CM, Golden JW
  TITLE     Regulation of gene expression in diverse cyanobacterial species by 
            using theophylline-responsive riboswitches.
  JOURNAL   Appl Environ Microbiol. 2014 Nov;80(21):6704-13. doi: 
            10.1128/AEM.01697-14. Epub 2014 Aug 22.
  PUBMED    25149516
REFERENCE   2  (bases 1 to 8164)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Dec 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; doi: 
            "10.1128/AEM.01697-14"; journalName: "Appl Environ Microbiol"; date:
            "2014-11"; volume: "80"; issue: "21"; pages: "6704-13"
COMMENT     Sequence Label: pAM5057
FEATURES             Location/Qualifiers
     source          1..8164
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             complement(397..1248)
                     /codon_start=1
                     /gene="repC"
                     /product="replication protein C of the broad-host-range 
                     plasmid RSF1010 (Scholz et al., 1989)"
                     /label=RSF1010 RepC
                     /translation="MVKPKNKHSLSHVRHDPAHCLAPGLFRALKRGERKRSKLDVTYDY
                     GDGKRIEFSGPEPLGADDLRILQGLVAMAGPNGLVLGPEPKTEGGRQLRLFLEPKWEAV
                     TADAMVVKGSYRALAKEIGAEVDSGGALKHIQDCIERLWKVSIIAQNGRKRQGFRLLSE
                     YASDEADGRLYVALNPLIAQAVMGGGQHVRISMDEVRALDSETARLLHQRLCGWIDPGK
                     TGKASIDTLCGYVWPSEASGSTMRKRRQRVREALPELVALGWTVTEFAAGKYDITRPKA
                     AG"
     CDS             complement(1235..2074)
                     /codon_start=1
                     /gene="repA"
                     /product="replication protein A of the broad-host-range 
                     plasmid RSF1010 (Scholz et al., 1989)"
                     /label=RSF1010 RepA
                     /translation="MATHKPINILEAFAAAPPPLDYVLPNMVAGTVGALVSPGGAGKSM
                     LALQLAAQIAGGPDLLEVGELPTGPVIYLPAEDPPTAIHHRLHALGAHLSAEERQAVAD
                     GLLIQPLIGSLPNIMAPEWFDGLKRAAEGRRLMVLDTLRRFHIEEENASGPMAQVIGRM
                     EAIAADTGCSIVFLHHASKGAAMMGAGDQQQASRGSSVLVDNIRWQSYLSSMTSAEAEE
                     WGVDDDQRRFFVRFGVSKANYGAPFADRWFRRHDGGVLKPAVLERQRKSKGVPRGEA"
     CDS             complement(2585..3556)
                     /codon_start=1
                     /gene="repB"
                     /product="replication protein B of the broad-host-range 
                     plasmid RSF1010 (Scholz et al., 1989)"
                     /label=RSF1010 RepB
                     /translation="MKNDRTLQAIGRQLKAMGCERFDIGVRDATTGQMMNREWSAAEVL
                     QNTPWLKRMNAQGNDVYIRPAEQERHGLVLVDDLSEFDLDDMKAEGREPALVVETSPKN
                     YQAWVKVADAAGGELRGQIARTLASEYDADPASADSRHYGRLAGFTNRKDKHTTRAGYQ
                     PWVLLRESKGKTATAGPALVQQAGQQIEQAQRQQEKARRLASLELSERQLSRHRRTALD
                     EYRSEMAGLVKRFGDDLSKCDFIAAQKLASRGRSAEEIGKAMAEASPALAERKPGHEAD
                     YIERTVSKVMGLPSVQLARAELARAPAPRQRGMDRGGPDFSM"
     oriT            4822..4909
                     /label=RSF1010 oriT
                     /note="origin of transfer of the broad-host-range plasmid 
                     RSF1010 (Scholz et al., 1989)"
     rep_origin      5251..5645
                     /label=RSF1010 oriV
                     /note="replication origin of the broad-host-range plasmid 
                     RSF1010; requires the RSF1010 RepA/B/C proteins for 
                     replication (Scholz et al., 1989)"
     terminator      complement(5808..5851)
                     /label=bacterial terminator
                     /note="putative bacterial transcription terminator"
     CDS             complement(5965..6753)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     promoter        join(7183..7188,7189..7204,7205..7211)
                     /label=tac promoter
                     /note="strong E. coli promoter; hybrid between the trp and 
                     lac UV5 promoters"
     CDS             7277..7306
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     protein_bind    7307..7331
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             join(7352..7354,7355..7357,7358..8071)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVRFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(7658..7677)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     8005..8026
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     terminator      8102..8129
                     /label=rrnB T2 terminator
                     /note="transcription terminator T2 from the E. coli rrnB 
                     gene"
ORIGIN
        1 tctagaacaa tggaagtcct cgtctcgccc tcgaattagc ccgcttaatg agcgggcttt
       61 ttttgaatta attctcgcga gctggcacga caggtttccc gactggaaag cgggcagtga
      121 gcgcaacgca attaatgtaa gttagcgcga attgcaagct ggccgacgcg ctgggctacg
      181 tcttgctggc gttcgggagc agaagagcat acatctggaa gcaaagccag gaaagcggcc
      241 tatggagctg tgcggcagcg ctcagtaggc aatttttcaa aatattgtta agccttttct
      301 gagcatggta tttttcatgg tattaccaat tagcaggaaa ataagccatt gaatataaaa
      361 gataaaaatg tcttgtttac aatagagtgg ggggggtcag cctgccgcct tgggccgggt
      421 gatgtcgtac ttgcccgccg cgaactcggt taccgtccag cccagcgcga ccagctccgg
      481 caacgcctcg cgcacccgct ggcggcgctt gcgcatggtc gaaccactgg cctctgacgg
      541 ccagacatag ccgcacaagg tatctatgga agccttgccg gttttgccgg ggtcgatcca
      601 gccacacagc cgctggtgca gcaggcgggc ggtttcgctg tccagcgccc gcacctcgtc
      661 catgctgatg cgcacatgct ggccgccacc catgacggcc tgcgcgatca aggggttcag
      721 ggccacgtac aggcgcccgt ccgcctcgtc gctggcgtac tccgacagca gccgaaaccc
      781 ctgccgcttg cggccattct gggcgatgat ggataccttc caaaggcgct cgatgcagtc
      841 ctgtatgtgc ttgagcgccc caccactatc gacctctgcc ccgatttcct ttgccagcgc
      901 ccgatagcta cctttgacca ccatggcatc agcggtgacg gcctcccact tgggttccag
      961 gaacagccgg agctgccgtc cgccttcggt cttgggttcc gggccaagca ctaggccatt
     1021 aggcccagcc atggccacca gcccttgcag gatgcgcaga tcatcagcgc ccagcggctc
     1081 cgggccgctg aactcgatcc gcttgccgtc gccgtagtca tacgtcacgt ccagcttgct
     1141 gcgcttgcgc tcgccccgct tgagggcacg gaacaggccg ggggccagac agtgcgccgg
     1201 gtcgtgccgg acgtggctga ggctgtgctt gttcttaggc ttcaccacgg ggcaccccct
     1261 tgctcttgcg ctgcctctcc agcacggcgg gcttgagcac cccgccgtca tgccgcctga
     1321 accaccgatc agcgaacggt gcgccatagt tggccttgct cacaccgaag cggacgaaga
     1381 accggcgctg gtcgtcgtcc acaccccatt cctcggcctc ggcgctggtc atgctcgaca
     1441 ggtaggactg ccagcggatg ttatcgacca gtaccgagct gccccggctg gcctgctgct
     1501 ggtcgcctgc gcccatcatg gccgcgccct tgctggcatg gtgcaggaac acgatagagc
     1561 acccggtatc ggcggcgatg gcctccatgc gaccgatgac ctgggccatg gggccgctgg
     1621 cgttttcttc ctcgatgtgg aaccggcgca gcgtgtccag caccatcagg cggcggccct
     1681 cggcggcgcg cttgaggccg tcgaaccact ccggggccat gatgttgggc aggctgccga
     1741 tcagcggctg gatcagcagg ccgtcagcca cggcttgccg ttcctcggcg ctgaggtgcg
     1801 ccccaagggc gtgcaggcgg tgatgaatgg cggtgggcgg gtcttcggcg ggcaggtaga
     1861 tcaccgggcc ggtgggcagt tcgcccacct ccagcagatc cggcccgcct gcaatctgtg
     1921 cggccagttg cagggccagc atggatttac cggcaccacc gggcgacacc agcgccccga
     1981 ccgtaccggc caccatgttg ggcaaaacgt agtccagcgg tggcggcgct gctgcgaacg
     2041 cctccagaat attgataggc ttatgggtag ccattgattg cctcctttgc aggcagttgg
     2101 tggttaggcg ctggcggggt cactaccccc gccctgcgcc gctctgagtt cttccaggca
     2161 ctcgcgcagc gcctcgtatt cgtcgtcggt cagccagaac ttgcgctgac gcatcccttt
     2221 ggccttcatg cgctcggcat atcgcgcttg gcgtacagcg tcagggctgg ccagcaggtc
     2281 gccggtctgc ttgtcctttt ggtctttcat atcagtcacc gagaaacttg ccggggccga
     2341 aaggcttgtc ttcgcggaac aaggacaagg tgcagccgtc aaggttaagg ctggccatat
     2401 cagcgactga aaagcggcca gcctcggcct tgtttgacgt ataaccaaag ccaccgggca
     2461 accaatagcc cttgtcactt ttgatcaggt agaccgaccc tgaagcgctt ttttcgtatt
     2521 ccataaaacc cccttctgtg cgtgagtact catagtataa caggcgtgag taccaacgca
     2581 agcactacat gctgaaatct ggcccgcccc tgtccatgcc tcgctggcgg ggtgccggtg
     2641 cccgtgccag ctcggcccgc gcaagctgga cgctgggcag acccatgacc ttgctgacgg
     2701 tgcgctcgat gtaatccgct tcgtggccgg gcttgcgctc tgccagcgct gggctggcct
     2761 cggccatggc cttgccgatt tcctcggcac tgcggccccg gctggccagc ttctgcgcgg
     2821 cgataaagtc gcacttgctg aggtcatcgc cgaagcgctt gaccagcccg gccatctcgc
     2881 tgcggtactc gtccagcgcc gtgcgccggt ggcggctaag ctgccgctcg gacagttcga
     2941 ggctggccag cctgcgggcc ttctcctgct gccgctgggc ctgctcgatc tgctggccag
     3001 cctgctgcac cagcgccggg ccagcggtgg cggtcttgcc cttggattca cgcagcagca
     3061 cccacggctg ataaccggcg cgggtggtgt gcttgtcctt gcggttggtg aagcccgcca
     3121 agcggccata gtggcggctg tcggcgctgg ccgggtcggc gtcgtactcg ctggccagcg
     3181 tccgggcaat ctgtccccga agttcaccgc ctgcggcgtc ggccaccttg acccatgcct
     3241 gatagttctt cgggctggtt tccactacca gggcaggctc ccggccctcg gctttcatgt
     3301 catccaggtc aaactcgctg aggtcgtcca ccagcaccag accatgccgc tcctgctcgg
     3361 cgggcctgat atacacgtca ttgccctggg cattcatccg cttgagccat ggcgtgttct
     3421 ggagcacttc ggcggctgac cattcccggt tcatcatctg gccggtggtg gcgtccctga
     3481 cgccgatatc gaagcgctca cagcccatgg ccttgagctg tcggcctatg gcctgcaaag
     3541 tcctgtcgtt cttcatcggg ccaccaagcg cagccagatc gagccgtcct cggttgtcag
     3601 tggcgtcagg tcgagcaaga gcaacgatgc gatcagcagc accaccgtag gcatcatgga
     3661 agccagcatc acggttagcc atagcttcca gtgccacccc cgcgacgcgc tccgggcgct
     3721 ctgcgcggcg ctgctcacct cggcggctac ctcccgcaac tctttggcca gctccaccca
     3781 tgccgcccct gtctggcgct gggctttcag ccactccgcc gcctgcgcct cgctggcctg
     3841 ctgggtctgg ctcatgacct gccgggcttc gtcggccagt gtcgccatgc tctgggccag
     3901 cggttcgatc tgctccgcta actcgttgat gcctctggat ttcttcactc tgtcgattgc
     3961 gttcatggtc tattgcctcc cggtattcct gtaagtcgat gatctgggcg ttggcggtgt
     4021 cgatgttcag ggccacgtct gcccggtcgg tgcggatgcc ccggccttcc atctccacca
     4081 cgttcggccc caggtgaaca ccgggcaggc gctcgatgcc ctgcgcctca agtgttctgt
     4141 ggtcaatgcg ggcgtcgtgg cctgcccgct ctaatgcccg gttggcatgg tcggcccatg
     4201 cctcgcgggt ctgctcaagc catgccttgg gcttgagcgc ttcggtcttc tgtgccccgc
     4261 ccttctccgg ggtcttgccg ttgtaccgct tgaaccactg agcggcgggc cgctcgatgc
     4321 cgtcattgat ccgctcggag atcatcaggt ggcagtgcgg gttctcgccg ccaccggcat
     4381 ggatggccag cgtatacggc aggcgctcgg caccggtcag gtgctgggcg aactcggacg
     4441 ccagcgcctt ctgctggtcg agggtcagct cgaccggcag ggcaaattcg acctccttga
     4501 acagccgccc attggcgcgt tcatacaggt cggcagcatc ccagtagtcg gcgggccgct
     4561 cgacgaactc cggcatgtgc ccggattcgg cgtgcaagac ttcatccatg tcgcgggcat
     4621 acttgccttc gcgctggatg aagtcggcct tgccttcgcg ctggatgaag tcggccttgg
     4681 ccctggccga ttggccgccc gacctgctgc cggttttcgc cgtaaggtga taaatcgcca
     4741 tgctgcctcg ctgttgcttt tgcttttcgg ctccatgcaa tggccctcgg agagcgcacc
     4801 gcccgaaggg tggccgttag gccagtttct cgaagagaaa ccggtaagtg cgccctcccc
     4861 tacaaagtag ggtcgggatt gccgccgctg tgcctccatg atagcctacg agacagcaca
     4921 ttaacaatgg ggtgtcaaga tggttaaggg gagcaacaag gcggcggatc ggctggccaa
     4981 gctcgaagaa caacgagcgc gaatcaatgc cgaaattcag cgggtgcggg caagggaaca
     5041 gcagcaagag cgcaagaacg aaacaaggcg caaggtgctg gtgggggcca tgattttggc
     5101 caaggtgaac agcagcgagt ggccggagga tcggctcatg gcggcaatgg atgcgtacct
     5161 tgaacgcgac cacgaccgcg ccttgttcgg tctgccgcca cgccagaagg atgagccggg
     5221 ctgaatgatc gaccgagaca ggccctgcgg ggctgcacac gcgcccccac ccttcgggta
     5281 gggggaaagg ccgctaaagc ggctaaaagc gctccagcgt atttctgcgg ggtttggtgt
     5341 ggggtttagc gggctttgcc cgcctttccc cctgccgcgc agcggtgggg cggtgtgtag
     5401 cctagcgcag cgaatagacc agctatccgg cctctggccg ggcatattgg gcaagggcag
     5461 cagcgcccca caagggcgct gataaccgcg cctagtggat tattcttaga taatcatgga
     5521 tggatttttc caacaccccg ccagcccccg cccctgctgg gtttgcaggt ttgggggcgt
     5581 gacagttatt gcaggggttc gtgacagtta ttgcaggggg gcgtgacagt tattgcaggg
     5641 gttcgtgaca gttagtacgg gagtgacggg cactggctgg caatgtctag caacggcagg
     5701 catttcggct gagggtaaaa gaactttccg ctaagcgata gactgtatgt aaacacagta
     5761 ttgcaaggac gcggaacatg cctcatgtgg cggccctcgc gagaattaat tcagataaaa
     5821 aaaatcctta gctttcgcta aggatgattt ctagcgatga ccctgctgat tggttcgctg
     5881 accgctagca tcgcgccgcg cgcgcgcgcg cggatgctag cggcgcgcca aaaaaaagcc
     5941 cgctcattag gcgggctggc cggccttgcc gactaccttg gtgatctcgc ctttcacgta
     6001 gtggacaaat tcttccaact gatctgcgcg cgaggccaag cgatcttctt cttgtccaag
     6061 ataagcctgt ctagcttcaa gtatgacggg ctgatactgg gccggcaggc gctccattgc
     6121 ccagtcggca gcgacatcct tcggcgcgat tttgccggtt actgcgctgt accaaatgcg
     6181 ggacaacgta agcactacat ttcgctcatc gccagcccag tcgggcggcg agttccatag
     6241 cgttaaggtt tcatttagcg cctcaaatag atcctgttca ggaaccggat caaagagttc
     6301 ctccgccgct ggacctacca aggcaacgct atgttctctt gcttttgtca gcaagatagc
     6361 cagatcaatg tcgatcgtgg ctggctcgaa gatacctgca agaatgtcat tgcgctgcca
     6421 ttctccaaat tgcagttcgc gcttagctgg ataacgccac ggaatgatgt cgtcgtgcac
     6481 aacaatggtg acttctacag cgcggagaat ctcgctctct ccaggggaag ccgaagtttc
     6541 caaaaggtcg ttgatcaaag ctcgccgcgt tgtttcatca agccttacgg tcaccgtaac
     6601 cagcaaatca atatcactgt gtggcttcag gccgccatcc actgcggagc cgtacaaatg
     6661 tacggccagc aacgtcggtt cgagatggcg ctcgatgacg ccaactacct ctgatagttg
     6721 agtcgatact tcggcgatca ccgcttccct catgatgttt aactttgttt tagggcgact
     6781 gccctgctgc gtaacatcgt tgctgctcca taacatcaaa catcgaccca cggcgtaacg
     6841 cgcttgctgc ttggatgccc gaggcataga ctgtacccca aaaaaacagt cataacaagc
     6901 catgaaaacc gccactgcgc cgttaccacc gctgcgttcg gtcaaggttc tggaccagtt
     6961 gcgtgagcgc atacgctact tgcattacag cttacgaacc gaacaggctt atgtccactg
     7021 ggttcgtgcc ttcatccgtt tccacggtgt gcgtcacccg gcaaccttgg gcagcagcga
     7081 agtcgaggca tttctgtcct ggctggcgaa cgagcgcaag gtttcggtct ccacgcattc
     7141 gaaaccggta tcccgccgcc gccgccggat accggtttcg aattgacaat taatcatcgg
     7201 ctcgtataat ggtaccggtg ataccagcat cgtcttgatg cccttggcag caccctgcta
     7261 aggacgcaac aagatggaac aaaaactgat tagcgaagaa gatctgacaa gtttgtacaa
     7321 aaaagcaggc tccgcggccg cccccttcac catggtgagc aagggcgagg agctgttcac
     7381 cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca agttcagcgt
     7441 gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt tcatctgcac
     7501 caccggcaag ctgcccgtgc cctggcccac cctcgtgacc accttcggct acggcctgca
     7561 gtgcttcgcc cgctaccccg accacatgaa gcagcacgac ttcttcaagt ccgccatgcc
     7621 cgaaggctac gtccaggagc gcaccatctt cttcaaggac gacggcaact acaagacccg
     7681 cgccgaggtg aggttcgagg gcgacaccct ggtgaaccgc atcgagctga agggcatcga
     7741 cttcaaggag gacggcaaca tcctggggca caagctggag tacaactaca acagccacaa
     7801 cgtctatatc atggccgaca agcagaagaa cggcatcaag gtgaacttca agatccgcca
     7861 caacatcgag gacggcagcg tgcagctcgc cgaccactac cagcagaaca cccccatcgg
     7921 cgacggcccc gtgctgctgc ccgacaacca ctacctgagc taccagtccg ccctgagcaa
     7981 agaccccaac gagaagcgcg atcacatggt cctgctggag ttcgtgaccg ccgccgggat
     8041 cactctcggc atggacgagc tgtacaagta acatgagccg tctcgacctc ggccctgcag
     8101 gagaaggcca tcctgacgga tggccttttt gaagctttct agaatcgggg gcccccgggg
     8161 ggat
//
