LOCUS       sequence-447740-        9093 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transcriptional unit for copper- and flipase-regulated BeYDV 
            Rep/RepA. OCSt, flanked by FRT sites, is embedded in the 5'UTR of 
            miniDFR..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9093)
  AUTHORS   Garcia-Perez E, Vazquez-Vilar M, Lozano-Duran R, Orzaez D
  TITLE     CuBe: a geminivirus-based copper-regulated expression system 
            suitable for post-harvest activation.
  JOURNAL   Plant Biotechnol J. 2025 Jan;23(1):141-155. doi: 10.1111/pbi.14485. 
            Epub 2024 Oct 22.
  PUBMED    39435699
REFERENCE   2  (bases 1 to 9093)
  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: "10.1111/pbi.14485";
            journalName: "Plant Biotechnol J"; date: "2025-01"; volume: "23"; 
            issue: "1"; pages: "141-155"
COMMENT     Sequence Label: pDGB3_alpha1_CBS-FRT-OCS-BeYDVRepRepA-tNos (GB...
FEATURES             Location/Qualifiers
     source          1..9093
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    315..348
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     terminator      349..1056
                     /label=OCS terminator
                     /note="octopine synthase terminator"
     protein_bind    1057..1090
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     terminator      2249..2501
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     primer_bind     complement(2536..2554)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     misc_feature    2742..2766
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     CDS             4067..4696
                     /codon_start=1
                     /product="stability protein from the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
                     /label=pVS1 StaA
                     /translation="MKVIAVLNQKGGSGKTTIATHLARALQLAGADVLLVDSDPQGSAR
                     DWAAVREDQPLTVVGIDRPTIDRDVKAIGRRDFVVIDGAPQAADLAVSAIKAADFVLIP
                     VQPSPYDIWATADLVELVKQRIEVTDGRLQAAFVVSRAIKGTRIGGEVAEALAGYELPI
                     LESRITQRVSYPGTAAAGTTVLESEPEGDAAREVQALAAEIKSKLI"
     CDS             5125..6198
                     /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      6264..6458
                     /label=pVS1 oriV
                     /note="origin of replication for the Pseudomonas plasmid 
                     pVS1 (Heeb et al., 2000)"
     primer_bind     complement(6694..6716)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     misc_feature    6802..6942
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     6857..6876
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     complement(6957..6974)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7128..7716)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7208..7227)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(7804..8619)
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     CDS             complement(7804..8619)
                     /codon_start=1
                     /gene="aph(3')-Ia"
                     /product="aminoglycoside phosphotransferase"
                     /label=KanR
                     /note="confers resistance to kanamycin in bacteria or G418 
                     (Geneticin(R)) in eukaryotes"
                     /translation="MSHIQRETSCSRPRLNSNMDADLYGYKWARDNVGQSGATIYRLYG
                     KPDAPELFLKHGKGSVANDVTDEMVRLNWLTEFMPLPTIKHFIRTPDDAWLLTTAIPGK
                     TAFQVLEEYPDSGENIVDALAVFLRRLHSIPVCNCPFNSDRVFRLAQAQSRMNNGLVDA
                     SDFDDERNGWPVEQVWKEMHKLLPFSPDSVVTHGDFSLDNLIFDEGKLIGCIDVGRVGI
                     ADRYQDLAILWNCLGEFSPSLQKRLFQKYGIDNPDMNKLQFHLMLDEFF"
     primer_bind     8527..8546
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     misc_feature    9052..9076
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA"
ORIGIN
        1 ggagagtggt cttgggagag cgatgcgtct tttccgatga accgttccag caaaaaagac
       61 taagcgatgc gtcttttccg ctgaaccgtt ccagcaaaaa agactaagcg atgcgtcttt
      121 tccgctgaac cgttccagca aaaaagacta ggagagcgat gcgtcttttc cgctgaaccg
      181 ttccagcaaa aaagactatc ccgtcctaat acctctacgt gcaagcagta taaaaacaaa
      241 atacataaac ataactaaaa tctcatatcg ttgtaattgt catatactag tggtaattaa
      301 gtccagttac acttgaagtt cctattctct agaaagtata ggaacttcct gctttaatga
      361 gatatgcgag aagcctatga tcgcatgata tttgctttca attctgttgt gcacgttgta
      421 aaaaacctga gcatgtgtag ctcagatcct taccgccggt ttcggttcat tctaatgaat
      481 atatcacccg ttactatcgt atttttatga ataatattct ccgttcaatt tactgattgt
      541 accctactac ttatatgtac aatattaaaa tgaaaacaat atattgtgct gaataggttt
      601 atagcgacat ctatgataga gcgccacaat aacaaacaat tgcgttttat tattacaaat
      661 ccaattttaa aaaaagcggc agaaccggtc aaacctaaaa gactgattac ataaatctta
      721 ttcaaatttc aaaagtgccc caggggctag tatctacgac acaccgagcg gcgaactaat
      781 aacgctcact gaagggaact ccggttcccc gccggcgcgc atgggtgaga ttccttgaag
      841 ttgagtattg gccgtccgct ctaccgaaag ttacgggcac cattcaaccc ggtccagcac
      901 ggcggccggg taaccgactt gctgccccga gaattatgca gcattttttt ggtgtatgtg
      961 ggccccaaat gaagtgcagg tcaaaccttg acagtgacga caaatcgttg ggcgggtcca
     1021 gggcgaattt tgcgacaaca tgtcgaggct cagcaggaag ttcctattct ctagaaagta
     1081 taggaacttc aaggttaaat ttgctgactc ttttttacct ttcatttctg aaaaatgcct
     1141 tctgctagta agaacttcag actccaatct aaatatgttt tccttaccta tcccaagtgc
     1201 tcatctcaaa gagatgattt attccagttt ctctgggaga aactcacacc ttttcttatt
     1261 ttcttccttg gtgttgcttc tgagcttcat caagatggca ctacccacta tcatgctctt
     1321 ctccagcttg ataaaaaacc ttgtattagg gatccttctt ttttcgattt tgaaggaaat
     1381 caccctaata tccagccagc tagaaactct aaacaagtcc ttgattacat atcaaaggac
     1441 ggagatatta aaaccagagg agatttccga gatcataagg tttctcctcg caaatctgac
     1501 gcacgatgga gaactattat ccagactgca acgtctaagg aggaatatct tgacatgatc
     1561 aaggaagaat tccctcatga atgggcaaca aagcttcaat ggctggaata ttcagccaac
     1621 aaattattcc ctccacaacc tgaaccgtat gtgtcgccct tcacagaatc agatcttcgc
     1681 tgccacgaag atctacactc ctggagggaa acccatctat accatgtaag catagacgct
     1741 tatacttaca tacatcctgt ctcataccaa caagctcaat ctgaccttga atggatggcc
     1801 gatttaacca ggacaatgga aggaatggaa tccgacaccc cagcctctac atctgcggac
     1861 caactcgtac cggaaagacc acctgggcta gaagtctcgg acgacacaac tattggaacg
     1921 gtaccatcga tttcaccaac tacgatgaac acgccaccta taatatcatc gacgacatcc
     1981 ccttcaagtt cgtcccattg tggaagcaat taataggttg ccagtctgat ttcactgtca
     2041 accctaaata tggaaaaaag aagaaaataa aaggtgggat cccttctata attctttgca
     2101 atcctgacga agactggatg ttatcaatga caagtcaaca gaaggattac tttaaagata
     2161 attgcgtcac ccactacatg tgtgacgggg agactttttt tgctcgggaa tcgtcgagtc
     2221 actgagcttg gaatggatct tcgatcccga tcgttcaaac atttggcaat aaagtttctt
     2281 aagattgaat cctgttgccg gtcttgcgac gattatcata taatttctgt tgaattacgt
     2341 taagcatgta ataattaaca tgtaatgcat gacgttattt atgagatggg tttttatgat
     2401 tagagtcccg caattataca tttaatacgc gatagaaaac aaaatatagc gcgcaaacta
     2461 ggataaatta tcgcgcgcgg tgtcatctat gttactagat cgggaattgc caagctaatt
     2521 cttgaagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc atgataataa
     2581 tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt
     2641 tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc
     2701 ttcaataatg ggaccgactc gcgctgtcat gagacgaatt ctgacaggat atattggcgg
     2761 gtaaacctaa gagaaaagag cgtttattag aataatcgga tatttaaaag ggcgtgaaaa
     2821 ggtttatccg ttcgtccatt tgtatgtgca tgccaaccac agggttcccc tcgggatcaa
     2881 agtactttga tccaacccct ccgctgctat agtgcagtcg gcttctgacg ttcagtgcag
     2941 ccgtcatctg aaaacgacat gtcgcacaag tcctaagtta cgcgacaggc tgccgccctg
     3001 cccttttcct ggcgttttct tgtcgcgtgt tttagtcgca taaagtagaa tacttgcgac
     3061 tagaaccgga gacattacgc catgaacaag agcgccgccg ctggcctgct gggctatgcc
     3121 cgcgtcagca ccgacgacca ggacttgacc aaccaacggg ccgaactgca cgcggccggc
     3181 tgcaccaagc tgttttccga gaagatcacc ggcaccaggc gcgaccgccc ggagctggcc
     3241 aggatgcttg accacctacg ccctggcgac gttgtgacag tgaccaggct agaccgcctg
     3301 gcccgcagca cccgcgacct actggacatt gccgagcgca tccaggaggc cggcgcgggc
     3361 ctgcgtagcc tggcagagcc gtgggccgac accaccacgc cggccggccg catggtgttg
     3421 accgtgttcg ccggcattgc cgagttcgag cgttccctaa tcatcgaccg cacccggagc
     3481 gggcgcgagg ccgccaaggc ccgaggcgtg aagtttggcc cccgccctac cctcaccccg
     3541 gcacagatcg cgcacgcccg cgagctgatc gaccaggaag gccgcaccgt gaaagaggcg
     3601 gctgcactgc ttggcgtgca tcgctcgacc ctgtaccgcg cacttgagcg cagcgaggaa
     3661 gtgacgccca ccgaggccag gcggcgcggt gccttccgtg aggacgcatt gaccgaggcc
     3721 gacgccctgg cggccgccga gaatgaacgc caagaggaac aagcatgaaa ccgcaccagg
     3781 acggccagga cgaaccgttt ttcattaccg aagagatcga ggcggagatg atcgcggccg
     3841 ggtacgtgtt cgagccgccc gcgcacctct caaccgtgcg gctgcatgaa atcctggccg
     3901 gtttgtctga tgccaagctg gcggcctggc cggccagctt ggccgctgaa gaaaccgagc
     3961 gccgccgtct aaaaaggtga tgtgtatttg agtaaaacag cttgcgtcat gcggtcgctg
     4021 cgtatatgat gcgatgagta aataaacaaa tacgcaaggg gaacgcatga aggttatcgc
     4081 tgtacttaac cagaaaggcg ggtcaggcaa gacgaccatc gcaacccatc tagcccgcgc
     4141 cctgcaactc gccggggccg atgttctgtt agtcgattcc gatccccagg gcagtgcccg
     4201 cgattgggcg gccgtgcggg aagatcaacc gctaaccgtt gtcggcatcg accgcccgac
     4261 gattgaccgc gacgtgaagg ccatcggccg gcgcgacttc gtagtgatcg acggagcgcc
     4321 ccaggcggcg gacttggctg tgtccgcgat caaggcagcc gacttcgtgc tgattccggt
     4381 gcagccaagc ccttacgaca tatgggccac cgccgacctg gtggagctgg ttaagcagcg
     4441 cattgaggtc acggatggaa ggctacaagc ggcctttgtc gtgtcgcggg cgatcaaagg
     4501 cacgcgcatc ggcggtgagg ttgccgaggc gctggccggg tacgagctgc ccattcttga
     4561 gtcccgtatc acgcagcgcg tgagctaccc aggcactgcc gccgccggca caaccgttct
     4621 tgaatcagaa cccgagggcg acgctgcccg cgaggtccag gcgctggccg ctgaaattaa
     4681 atcaaaactc atttgagtta atgaggtaaa gagaaaatga gcaaaagcac aaacacgcta
     4741 agtgccggcc gtccgagcgc acgcagcagc aaggctgcaa cgttggccag cctggcagac
     4801 acgccagcca tgaagcgggt caactttcag ttgccggcgg aggatcacac caagctgaag
     4861 atgtacgcgg tacgccaagg caagaccatt accgagctgc tatctgaata catcgcgcag
     4921 ctaccagagt aaatgagcaa atgaataaat gagtagatga attttagcgg ctaaaggagg
     4981 cggcatggaa aatcaagaac aaccaggcac cgacgccgtg gaatgcccca tgtgtggagg
     5041 aacgggcggt tggccaggcg taagcggctg ggttgcctgc cggccctgca atggcactgg
     5101 aacccccaag cccgaggaat cggcgtgagc ggtcgcaaac catccggccc ggtacaaatc
     5161 ggcgcggcgc tgggtgatga cctggtggag aagttgaagg ccgcgcaggc cgcccagcgg
     5221 caacgcatcg aggcagaagc acgccccggt gaatcgtggc aagcggccgc tgatcgaatc
     5281 cgcaaagaat cccggcaacc gccggcagcc ggtgcgccgt cgattaggaa gccgcccaag
     5341 ggcgacgagc aaccagattt tttcgttccg atgctctatg acgtgggcac ccgcgatagt
     5401 cgcagcatca tggacgtggc cgttttccgt ctgtcgaagc gtgaccgacg agctggcgag
     5461 gtgatccgct acgagcttcc agacgggcac gtagaggttt ccgcagggcc ggccggcatg
     5521 gccagtgtgt gggattacga cctggtactg atggcggttt cccatctaac cgaatccatg
     5581 aaccgatacc gggaagggaa gggagacaag cccggccgcg tgttccgtcc acacgttgcg
     5641 gacgtactca agttctgccg gcgagccgat ggcggaaagc agaaagacga cctggtagaa
     5701 acctgcattc ggttaaacac cacgcacgtt gccatgcagc gtacgaagaa ggccaagaac
     5761 ggccgcctgg tgacggtatc cgagggtgaa gccttgatta gccgctacaa gatcgtaaag
     5821 agcgaaaccg ggcggccgga gtacatcgag atcgagctag ctgattggat gtaccgcgag
     5881 atcacagaag gcaagaaccc ggacgtgctg acggttcacc ccgattactt tttgatcgat
     5941 cccggcatcg gccgttttct ctaccgcctg gcacgccgcg ccgcaggcaa ggcagaagcc
     6001 agatggttgt tcaagacgat ctacgaacgc agtggcagcg ccggagagtt caagaagttc
     6061 tgtttcaccg tgcgcaagct gatcgggtca aatgacctgc cggagtacga tttgaaggag
     6121 gaggcggggc aggctggccc gatcctagtc atgcgctacc gcaacctgat cgagggcgaa
     6181 gcatccgccg gttcctaatg tacggagcag atgctagggc aaattgccct agcaggggaa
     6241 aaaggtcgaa aaggactctt tcctgtggat agcacgtaca ttgggaaccc aaagccgtac
     6301 attgggaacc ggaacccgta cattgggaac ccaaagccgt acattgggaa ccggtcacac
     6361 atgtaagtga ctgatataaa agagaaaaaa ggcgattttt ccgcctaaaa ctctttaaaa
     6421 cttattaaaa ctcttaaaac ccgcctggcc tgtgcataac tgtctggcca gcgcacagcc
     6481 gaagagctgc aaaaagcgcc tacccttcgg tcgctgcgct ccctacgccc cgccgcttcg
     6541 cgtcggccta tcgcggccgc tggccgctca aaaatggctg gcctacggcc aggcaatcta
     6601 ccagggcgcg gacaagccgc gccgtcgcca ctcgaccgcc ggcgcccaca tcaaggcacc
     6661 ctgcctcgcg cgtttcggtg atgacggtga aaacctctga cacatgcagc tcccggtgac
     6721 ggtcacagct tgtctgtaag cggatgccgg gagcagacaa gcccgtcagg gcgcgtcagc
     6781 gggtgttggc gggtgtcggg gcgcagccat gacccagtca cgtagcgata gcggagtgta
     6841 tactggctta actatgcggc atcagagcag attgtactga gagtgcacca tatgcggtgt
     6901 gaaataccgc acagatgcgt aaggagaaaa taccgcatca ggcgctcttc cgcttcctcg
     6961 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     7021 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     7081 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     7141 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     7201 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     7261 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     7321 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     7381 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     7441 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     7501 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     7561 actagaagga cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     7621 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     7681 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     7741 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gcattctagg
     7801 tgattagaaa aactcatcga gcatcaaatg aaactgcaat ttattcatat caggattatc
     7861 aataccatat ttttgaaaaa gccgtttctg taatgaagga gaaaactcac cgaggcagtt
     7921 ccataggatg gcaagatcct ggtatcggtc tgcgattccg actcgtccaa catcaataca
     7981 acctattaat ttcccctcgt caaaaataag gttatcaagt gagaaatcac catgagtgac
     8041 gactgaatcc ggtgagaatg gcaaaagttt atgcatttct ttccagactt gttcaacagg
     8101 ccagccatta cgctcgtcat caaaatcact cgcatcaacc aaaccgttat tcattcgtga
     8161 ttgcgcctga gcgagtcgaa atacgcgatc gctgttaaaa ggacaattac aaacaggaat
     8221 cgaatgcaac cggcgcagga acactgccag cgcatcaaca atattttcac ctgaatcagg
     8281 atattcttct aatacctgga atgctgtttt ccctgggatc gcagtggtga gtaaccatgc
     8341 atcatcagga gtacggataa aatgcttgat ggtcggaaga ggcataaatt ccgtcagcca
     8401 gtttagtctg accatctcat ctgtaacatc attggcaacg ctacctttgc catgtttcag
     8461 aaacaactct ggcgcatcgg gcttcccata caatcggtag attgtcgcac ctgattgccc
     8521 gacattatcg cgagcccatt tatacccata taaatcagca tccatgttgg aatttaatcg
     8581 cggccttgag caagacgttt cccgttgaat atggctcata acagaactta ttatttcctt
     8641 cctcttttct acagtattta aagatacccc aagaagctaa ttataacaag acgaactcca
     8701 attcactgtt ccttgcattc taaaacctta aataccagaa aacagctttt tcaaagttgt
     8761 tttcaaagtt ggcgtataac atagtatcga cggagccgat tttgaaaccg cggtgatcac
     8821 aggcagcaac gctctgtcat cgttacaatc aacatgctac cctccgcgag atcatccgtg
     8881 tttcaaaccc ggcagcttag ttgccgttct tccgaatagc atcggtaaca tgagcaaagt
     8941 ctgccgcctt acaacggctc tcccgctgac gccgtcccgg actgatgggc tgcctgtatc
     9001 gagtggtgat tttgtgccga gctgccggtc ggggagctgt tggctggctg gtggcaggat
     9061 atattgtggt gtaaacataa cgaattcgtc tca
//
