LOCUS       sequence-84577-e        8260 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8260)
  AUTHORS   Binder A, Lambert J, Morbitzer R, Popp C, Ott T, Lahaye T, Parniske 
            M
  TITLE     A modular plasmid assembly kit for multigene expression, gene 
            silencing and silencing rescue in plants.
  JOURNAL   PLoS One. 2014 Feb 13;9(2):e88218. doi: 
            10.1371/journal.pone.0088218. eCollection 2014.
  PUBMED    24551083
REFERENCE   2  (bases 1 to 8260)
  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"; journalName: "PLoS One.";
            date: "2014-02-13"; volume: "9(2):e88218. doi"; pages: " 
            10.1371/journal.pone.0088218. eCollection 2014"
COMMENT     Sequence Label: LIIbeta R 1-2
FEATURES             Location/Qualifiers
     source          1..8260
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     CDS             371..1162
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MGEAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
     misc_feature    1540..1564
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
     promoter        join(1758..1763,1764..1781,1782..1788)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             1842..2501
                     /codon_start=1
                     /gene="cat"
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQGGA"
     primer_bind     complement(1908..1927)
                     /label=CAT-R
                     /note="Chloramphenicol resistance gene, reverse primer"
     CDS             2843..3148
                     /codon_start=1
                     /gene="ccdB"
                     /product="CcdB, a bacterial toxin that poisons DNA gyrase"
                     /label=ccdB
                     /note="Plasmids containing the ccdB gene cannot be 
                     propagated in standard E. coli strains."
                     /translation="MQFKVYTYKRESRYRLFVDVQSDIIDTPGRRMVIPLASARLLSDK
                     VPRELYPVVHIGDESWRMMTTDMASVPVSVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     3027..3046
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     misc_feature    3234..3258
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             4559..5188
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
     CDS             5617..6690
                     /codon_start=1
                     /product="replication protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 RepA
                     /translation="MSGRKPSGPVQIGAALGDDLVEKLKAAQAAQRQRIEAEARPGESW
                     QAAADRIRKESRQPPAAGAPSIRKPPKGDEQPDFFVPMLYDVGTRDSRSIMDVAVFRLS
                     KRDRRAGEVIRYELPDGHVEVSAGPAGMASVWDYDLVLMAVSHLTESMNRYREGKGDKP
                     GRVFRPHVADVLKFCRRADGGKQKDDLVETCIRLNTTHVAMQRTKKAKNGRLVTVSEGE
                     ALISRYKIVKSETGRPEYIEIELADWMYREITEGKNPDVLTVHPDYFLIDPGIGRFLYR
                     LARRAAGKAEARWLFKTIYERSGSAGEFKKFCFTVRKLIGSNDLPEYDLKEEAGQAGPI
                     LVMRYRNLIEGEASAGS"
     rep_origin      6756..6950
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     primer_bind     complement(7186..7208)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    7294..7434
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     7349..7368
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(7449..7466)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7620..8208)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7700..7719)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 actaagctgc cgggtttgaa acacggatga tctcgcggag ggtagcatgt tgattgtaac
       61 gatgacagag cgttgctgcc tgtgatcaat tcgggcacga acccagtgga cataagcctg
      121 ttcggttcgt aagctgtaat gcaagtagcg tatgcgctca cgcaactggt ccagaacctt
      181 gaccgaacgc agcggtggta acggcgcagt ggcggttttc atggcttgtt atgactgttt
      241 ttttggggta cagtctatgc ctcgggcatc caagcagcaa gcgcgttacg ccgtgggtcg
      301 atgtttgatg ttatggagca gcaacgatgt tacgcagcag ggcagtcgcc ctaaaacaaa
      361 gttaaacatc atgggggaag cggtgatcgc cgaagtatcg actcaactat cagaggtagt
      421 tggcgtcatc gagcgccatc tcgaaccgac gttgctggcc gtacatttgt acggctccgc
      481 agtggatggc ggcctgaagc cacacagtga tattgatttg ctggttacgg tgaccgtaag
      541 gcttgatgaa acaacgcggc gagctttgat caacgacctt ttggaaactt cggcttcccc
      601 tggagagagc gagattctcc gcgctgtaga agtcaccatt gttgtgcacg acgacatcat
      661 tccgtggcgt tatccagcta agcgcgaact gcaatttgga gaatggcagc gcaatgacat
      721 tcttgcaggt atcttcgagc cagccacgat cgacattgat ctggctatct tgctgacaaa
      781 agcaagagaa catagcgttg ccttggtagg tccagcggcg gaggaactct ttgatccggt
      841 tcctgaacag gatctatttg aggcgctaaa tgaaacctta acgctatgga actcgccgcc
      901 cgactgggct ggcgatgagc gaaatgtagt gcttacgttg tcccgcattt ggtacagcgc
      961 agtaaccggc aaaatcgcgc cgaaggatgt cgctgccgac tgggcaatgg agcgcctgcc
     1021 ggcccagtat cagcccgtca tacttgaagc tagacaggct tatcttggac aagaagaaga
     1081 tcgcttggcc tcgcgcgcag atcagttgga agaatttgtc cactacgtga aaggcgagat
     1141 caccaaggta gtcggcaaat aatgtctagc tagaaattcg ttcaagccga cgccgcttcg
     1201 cggcgcggct taactcaagc gttagatgca ctaagcacat aattgctcac agccaaacta
     1261 tcaggtcaag tctgctttta ttatttttaa gcgtgcataa taagccctac acaaattggg
     1321 agatatatca tgcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     1381 ccccgtagaa aagatcaaag gatcttctgc cggtaacatg agcaaagtct gccgccttac
     1441 aacggctctc ccgctgacgc cgtcccggac tgatgggctg cctgtatcga gtggtgattt
     1501 tgtgccgagc tgccggtcgg ggagctgttg gctggctggt ggcaggatat attgtggtgt
     1561 aaacaaattg acgcttagac ggattacgga agacaatacg gacatgagac caaagctgaa
     1621 cgagaaacgt aaaatgatat aaatatcaat atattaaatt agattttgca taaaaaacag
     1681 actacataat actgtaaaac acaacatatc cagtcactat ggcggcccct cgaggggccg
     1741 cattaggcac cccaggcttt acactttatg cttccggctc gtataatgtg tggattttga
     1801 gttaggatcc gtcgagattt tcaggagcta aggaagctaa aatggagaaa aaaatcactg
     1861 gatataccac cgttgatata tcccaatggc atcgtaaaga acattttgag gcatttcagt
     1921 cagttgctca atgtacctat aaccagaccg ttcagctgga tattacggcc tttttaaaga
     1981 ccgtaaagaa aaataagcac aagttttatc cggcctttat tcacattctt gcccgcctga
     2041 tgaatgctca tccggaattc cgtatggcaa tgaaagacgg tgagctggtg atatgggata
     2101 gtgttcaccc ttgttacacc gttttccatg agcaaactga aacgttttca tcgctctgga
     2161 gtgaatacca cgacgatttc cggcagtttc tacacatata ttcgcaagat gtggcgtgtt
     2221 acggtgaaaa cctggcctat ttccctaaag ggtttattga gaatatgttt ttcgtgtcag
     2281 ccaatccctg ggtgagtttc accagttttg atttaaacgt ggccaatatg gacaacttct
     2341 tcgcccccgt tttcaccatg ggcaaatatt atacgcaagg cgacaaggtg ctgatgccgc
     2401 tggcgattca ggttcatcat gccgtttgtg atggcttcca tgtcggcaga atgcttaatg
     2461 aattacaaca gtactgcgat gagtggcagg gcggggcgta atctagagga tccggcttac
     2521 taaaagccag ataacagtat gcgtatttgc gcgctgattt ttgcggtata agaatatata
     2581 ctgatatgta tacccgaagt atgtcaaaaa gaggtatgct atgaagcagc gtattacagt
     2641 gacagttgac agcgacagct atcagttgct caaggcatat atgatgtcaa tatctccggt
     2701 ctggtaagca caaccatgca gaatgaagcc cgtcgtctgc gtgccgaacg ctggaaagcg
     2761 gaaaatcagg aagggatggc tgaggtcgcc cggtttattg aaatgaacgg ctcttttgct
     2821 gacgagaaca ggggctggtg aaatgcagtt taaggtttac acctataaaa gagagagccg
     2881 ttatcgtctg tttgtggatg tacagagtga tattattgac acgcccgggc gacggatggt
     2941 gatccccctg gccagtgcac gtctgctgtc agataaagtc ccccgtgaac tttacccggt
     3001 ggtgcatatc ggggatgaaa gctggcgcat gatgaccacc gatatggcca gtgtgccggt
     3061 ttccgttatc ggggaagaag tggctgatct cagccaccgc gaaaatgaca tcaaaaacgc
     3121 cattaacctg atgttctggg gaatataaat gtcaggctcc cttatacaca gccagtctgc
     3181 aggtcgacgg tctcaccgcg ggcttgtctt ctctggttta aactatcagt gtttgacagg
     3241 atatattggc gggtaaacct aagagaaaag agcgtttatt agaataatcg gatatttaaa
     3301 agggcgtgaa aaggtttatc cgttcgtcca tttgtatgtg catgccaacc acagggttcc
     3361 cctcgggatc aaagtacttt gatccaaccc ctccgctgct atagtgcagt cggcttctga
     3421 cgttcagtgc agccttcttc tgaaaacgac atgtcgcaca agtcctaagt tacgcgacag
     3481 gctgccgccc tgcccttttc ctggcgtttt cttgtcgcgt gttttagtcg cataaagtag
     3541 aatacttgcg actagaaccg gagacattac gccatgaaca agagcgccgc cgctggcctg
     3601 ctgggctatg cccgcgtcag caccgacgac caggacttga ccaaccaacg ggccgaactg
     3661 cacgcggccg gctgcaccaa gctgttttcc gagaagatca ccggcaccag gcgcgaccgc
     3721 ccggagctgg ccaggatgct tgaccaccta cgccctggcg acgttgtgac agtgaccagg
     3781 ctagaccgcc tggcccgcag cacccgcgac ctactggaca ttgccgagcg catccaggag
     3841 gccggcgcgg gcctgcgtag cctggcagag ccgtgggccg acaccaccac gccggccggc
     3901 cgcatggtgt tgaccgtgtt cgccggcatt gccgagttcg agcgttccct aatcatcgac
     3961 cgcacccgga gcgggcgcga ggccgccaag gcccgaggcg tgaagtttgg cccccgccct
     4021 accctcaccc cggcacagat cgcgcacgcc cgcgagctga tcgaccagga aggccgcacc
     4081 gtgaaagagg cggctgcact gcttggcgtg catcgctcga ccctgtaccg cgcacttgag
     4141 cgcagcgagg aagtgacgcc caccgaggcc aggcggcgcg gtgccttccg tgaggacgca
     4201 ttgaccgagg ccgacgccct ggcggccgcc gagaatgaac gccaagagga acaagcatga
     4261 aaccgcacca ggacggccag gacgaaccgt ttttcattac cgaagagatc gaggcggaga
     4321 tgatcgcggc cgggtacgtg ttcgagccgc ccgcgcacat ctcaaccgtg cggctgcatg
     4381 aaatcctggc cggtttgtct gatgccaagc tggcggcctg gccggccagc ttggccgctg
     4441 aagaaaccga gcgccgccgt ctaaaaaggt gatgtgtatt tgagtaaaac agcttgcgtc
     4501 atgcggtcgc tgcgtatatg atgcgatgag taaataaaca aatacgcaag gggaacgcat
     4561 gaaggttatc gctgtactta accagaaagg cgggtcaggc aagacgacca tcgcaaccca
     4621 tctagcccgc gccctgcaac tcgccggggc cgatgttctg ttagtcgatt ccgatcccca
     4681 gggcagtgcc cgcgattggg cggccgtgcg ggaagatcaa ccgctaaccg ttgtcggcat
     4741 cgaccgcccg acgattgacc gcgacgtgaa ggccatcggc cggcgcgact tcgtagtgat
     4801 cgacggagcg ccccaggcgg cggacttggc tgtgtccgcg atcaaggcag ccgacttcgt
     4861 gctgattccg gtgcagccaa gcccttacga catatgggcc accgccgacc tggtggagct
     4921 ggttaagcag cgcattgagg tcacggatgg aaggctacaa gcggcctttg tcgtgtcgcg
     4981 ggcgatcaaa ggcacgcgca tcggcggtga ggttgccgag gcgctggccg ggtacgagct
     5041 gcccattctt gagtcccgta tcacgcagcg cgtgagctac ccaggcactg ccgccgccgg
     5101 cacaaccgtt cttgaatcag aacccgaggg cgacgctgcc cgcgaggtcc aggcgctggc
     5161 cgctgaaatt aaatcaaaac tcatttgagt taatgaggta aagagaaaat gagcaaaagc
     5221 acaaacacgc taagtgccgg ccgtccgagc gcacgcagca gcaaggctgc aacgttggcc
     5281 agcctggcag acacgccagc catgaagcgg gtcaactttc agttgccggc ggaggatcac
     5341 accaagctga agatgtacgc ggtacgccaa ggcaagacca ttaccgagct gctatctgaa
     5401 tacatcgcgc agctaccaga gtaaatgagc aaatgaataa atgagtagat gaattttagc
     5461 ggctaaagga ggcggcatgg aaaatcaaga acaaccaggc accgacgccg tggaatgccc
     5521 catgtgtgga ggaacgggcg gttggccagg cgtaagcggc tgggttgcct gccggccctg
     5581 caatggcact ggaaccccca agcccgagga atcggcgtga gcggtcgcaa accatccggc
     5641 ccggtacaaa tcggcgcggc gctgggtgat gacctggtgg agaagttgaa ggccgcgcag
     5701 gccgcccagc ggcaacgcat cgaggcagaa gcacgccccg gtgaatcgtg gcaagcggcc
     5761 gctgatcgaa tccgcaaaga atcccggcaa ccgccggcag ccggtgcgcc gtcgattagg
     5821 aagccgccca agggcgacga gcaaccagat tttttcgttc cgatgctcta tgacgtgggc
     5881 acccgcgata gtcgcagcat catggacgtg gccgttttcc gtctgtcgaa gcgtgaccga
     5941 cgagctggcg aggtgatccg ctacgagctt ccagacgggc acgtagaggt ttccgcaggg
     6001 ccggccggca tggccagtgt gtgggattac gacctggtac tgatggcggt ttcccatcta
     6061 accgaatcca tgaaccgata ccgggaaggg aagggagaca agcccggccg cgtgttccgt
     6121 ccacacgttg cggacgtact caagttctgc cggcgagccg atggcggaaa gcagaaagac
     6181 gacctggtag aaacctgcat tcggttaaac accacgcacg ttgccatgca gcgtacgaag
     6241 aaggccaaga acggccgcct ggtgacggta tccgagggtg aagccttgat tagccgctac
     6301 aagatcgtaa agagcgaaac cgggcggccg gagtacatcg agatcgagct agctgattgg
     6361 atgtaccgcg agatcacaga aggcaagaac ccggacgtgc tgacggttca ccccgattac
     6421 tttttgatcg atcccggcat cggccgtttt ctctaccgcc tggcacgccg cgccgcaggc
     6481 aaggcagaag ccagatggtt gttcaagacg atctacgaac gcagtggcag cgccggagag
     6541 ttcaagaagt tctgtttcac cgtgcgcaag ctgatcgggt caaatgacct gccggagtac
     6601 gatttgaagg aggaggcggg gcaggctggc ccgatcctag tcatgcgcta ccgcaacctg
     6661 atcgagggcg aagcatccgc cggttcctaa tgtacggagc agatgctagg gcaaattgcc
     6721 ctagcagggg aaaaaggtcg aaaaggactc tttcctgtgg atagcacgta cattgggaac
     6781 ccaaagccgt acattgggaa ccggaacccg tacattggga acccaaagcc gtacattggg
     6841 aaccggtcac acatgtaagt gactgatata aaagagaaaa aaggcgattt ttccgcctaa
     6901 aactctttaa aacttattaa aactcttaaa acccgcctgg cctgtgcata actgtctggc
     6961 cagcgcacag ccgaagagct gcaaaaagcg cctacccttc ggtcgctgcg ctccctacgc
     7021 cccgccgctt cgcgtcggcc tatcgcggcc gctggccgct caaaaatggc tggcctacgg
     7081 ccaggcaatc taccagggcg cggacaagcc gcgccgtcgc cactcgaccg ccggcgccca
     7141 catcaaggca ccctgcctcg cgcgtttcgg tgatgacggt gaaaacctct gacacatgca
     7201 gctcccggag acagtcacag cttgtctgta agcggatgcc gggagcagac aagcccgtca
     7261 gggcgcgtca gcgggtgttg gcgggtgtcg gggcgcagcc atgacccagt cacgtagcga
     7321 tagcggagtg tatactggct taactatgcg gcatcagagc agattgtact gagagtgcac
     7381 catatgcggt gtgaaatacc gcacagatgc gtaaggagaa aataccgcat caggcgctct
     7441 tccgcttcct cgctcactga ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca
     7501 gctcactcaa aggcggtaat acggttatcc acagaatcag gggataacgc aggaaagaac
     7561 atgtgagcaa aaggccagca aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt
     7621 ttccataggc tccgcccccc tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg
     7681 cgaaacccga caggactata aagataccag gcgtttcccc ctggaagctc cctcgtgcgc
     7741 tctcctgttc cgaccctgcc gcttaccgga tacctgtccg cctttctccc ttcgggaagc
     7801 gtggcgcttt ctcatagctc acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc
     7861 aagctgggct gtgtgcacga accccccgtt cagcccgacc gctgcgcctt atccggtaac
     7921 tatcgtcttg agtccaaccc ggtaagacac gacttatcgc cactggcagc agccactggt
     7981 aacaggatta gcagagcgag gtatgtaggc ggtgctacag agttcttgaa gtggtggcct
     8041 aactacggct acactagaag gacagtattt ggtatctgcg ctctgctgaa gccagttacc
     8101 ttcggaaaaa gagttggtag ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt
     8161 ttttttgttt gcaagcagca gattacgcgc agaaaaaaag gatctcaaga agatcctttg
     8221 atcttttcta cggggtctga cgctcatcgg aagaacggca
//
