LOCUS       sequence-234600-        6199 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Binary expression vector containing pSUC2:MS2-VP64:tRBCS followed by
            GreenGate A-G sequences for insertion of additional modules.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6199)
  AUTHORS   Lee JE, Neumann M, Duro DI, Schmid M
  TITLE     CRISPR-based tools for targeted transcriptional and epigenetic 
            regulation in plants.
  JOURNAL   PLoS One. 2019 Sep 26;14(9):e0222778. doi: 
            10.1371/journal.pone.0222778. eCollection 2019.
  PUBMED    31557222
REFERENCE   2  (bases 1 to 6199)
  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: "2019-09-26"; volume: "14(9):e0222778. doi"; pages: " 
            10.1371/journal.pone.0222778. eCollection 2019"
COMMENT     Sequence Label: pMH205
FEATURES             Location/Qualifiers
     source          1..6199
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    534..556
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA 
                     (truncated)"
     CDS             2811..3197
                     /codon_start=1
                     /product="bacteriophage MS2 coat protein"
                     /label=MS2-N55K
                     /note="binds with high affinity to a specific stem-loop 
                     structure in the viral RNA (Lim et al., 1994)"
                     /translation="ASNFTQFVLVDNGGTGDVTVAPSNFANGVAEWISSNSRSQAYKVT
                     CSVRQSSAQKRKYTIKVEVPKVATQTVGGVELPVAAWRSYLNMELTIPIFATNSDCELI
                     VKAMQGLLKDGNPIPSAIAANSGIY"
     CDS             3252..3272
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3303..3452
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     terminator      3471..4112
                     /label=E9 terminator
                     /note="terminator and polyadenylation signal from the pea 
                     rbcS-E9 gene"
     misc_feature    4218..4242
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     rep_origin      complement(4333..4921)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4413..4432)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(5092..5880)
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEDRLASRADQLEEFVHYVKGEITKVVGK"
     rep_origin      join(6171..6199,1..407)
                     /label=pSa ori
                     /note="origin of replication from bacterial plasmid pSa"
ORIGIN
        1 tttttatccc cggaagcctg tggatagagg gtagttatcc acgtgaaacc gctaatgccc
       61 cgcaaagcct tgattcacgg ggctttccgg cccgctccaa aaactatcca cgtgaaatcg
      121 ctaatcaggg tacgtgaaat cgctaatcgg agtacgtgaa atcgctaata aggtcacgtg
      181 aaatcgctaa tcaaaaaggc acgtgagaac gctaatagcc ctttcagatc aacagcttgc
      241 aaacacccct cgctccggca agtagttaca gcaagtagta tgttcaatta gcttttcaat
      301 tatgaatata tatatcaatt attggtcgcc cttggcttgt ggacaatgcg ctacgcgcac
      361 cggctccgcc cgtggacaac cgcaagcggt tgcccaccgt cgagcgcctt tgcccacaac
      421 ccggcggccg gccgcaacag atcgttttat aaattttttt ttttgaaaaa gaaaaagccc
      481 gaaaggcggc aacctctcgg gcttctggat ttccgatccc cggaattaga tcttggcagg
      541 atatattgtg gtgtaacgtt ctcgaactag gctcggacga agtaagcggc cgcaacctta
      601 attaaacacg gcgcgccaac agcgatcgca ccaggtacca cctctaaaat ctggtttcat
      661 attaatttcg cacaccaagt tactttctat tattaactgt tataatggac catgaaatca
      721 tttgcatatg aactgcaatg atacataatc cactttgttt tgtgggagac atttaccaga
      781 tttcggtaaa ttggtattcc cccttttatg tgattggtca ttgatcattg ttagtggcca
      841 gacatttgaa ctcccgtttt tttgtctata agaattcgga aacatatagt atcctttgaa
      901 aacggagaaa caaataacaa tgtggacaaa ctagatataa tttcaacaca agactatggg
      961 aatgatttta cccactaatt ataatccgat cacaaggttt caacgaacta gttttccaga
     1021 tatcaaccaa atttactttg gaattaaact aacttaaaac taattggttg ttcgtaaatg
     1081 gtgctttttt tttttgcgga tgttagtaaa gggttttatg tattttatat tattagttat
     1141 ctgttttcag tgttatgttg tctcatccat aaagtttata tgttttttct ttgctctata
     1201 acttatatat atatatgagt ttacagttat atttatacat ttcagatact tgatcggcat
     1261 tttttttggt aaaaaatata tgcatgaaaa actcaagtgt ttctttttta aggaattttt
     1321 aaatggtgat tatatgaata taatcatatg tatatccgta tatatatgta gccagatagt
     1381 taattatttg ggggatattt gaattattaa tgttataata ttctttcttt tgactcgtct
     1441 ggttaaatta aagaacaaaa aaaacacata cttttactgt tttaaaaggt taaattaaca
     1501 taatttattg attacaagtg tcaagtccat gacattgcat gtaggttcga gacttcagag
     1561 ataacggaag agatcgataa ttgtgatcgt aacatccaga tatgtatgtt taattttcat
     1621 ttagatgtgg atcagagaag ataagtcaaa ctgtcttcat aatttaagac aacctctttt
     1681 aatattttcc caaaacatgt tttatgtaac tactttgctt atgtgattgc ctgaggatac
     1741 tattattctc tgtctttatt ctcttcacac cacatttaaa tagtttaaga gcatagaaat
     1801 taattatttt caaaaaggtg attatatgca tgcaaaatag cacaccattt atgtttatat
     1861 tttcaaatta tttaatacat ttcaatattt cataagtgtg attttttttt ttattgtcaa
     1921 tttcataagt gtgatttgtc atttgtatta aacaattgta tcgcgcagta caaataaaca
     1981 gtgggagagg tgaaaatgca gttataaaac tgtccaataa tttactaaca catttaaata
     2041 tctaaaaaga gtgtttcaaa aaaaattctt ttgaaataag aaaagtgata gatattttta
     2101 cgctttcgtc tgaaaataaa acaataatag tttattagaa aaatgttatc accgaaaatt
     2161 attctagtgc cactcgctcg gatcgaaatt cgaaagttat attctttctc tttacctaat
     2221 ataaaaatca caagaaaaat caatccgaat atatctatca acatagtata tgcccttaca
     2281 tattgtttct gacttttctc tatccgaatt tctcgcttca tggttttttt ttaacatatt
     2341 ctcatttaat tttcattact attatataac taaaagatgg aaataaaata aagtgtcttt
     2401 gagaatcgaa cgtccatatc agtaagatag tttgtgtgaa ggtaaaatct aaaagattta
     2461 agttccaaaa acagaaaata atatattacg ctaaaaaaga agaaaataat taaatacaaa
     2521 acagaaaaaa ataatatacg acagacacgt gtcacgaaga taccctacgc tatagacaca
     2581 gctctgtttt ctcttttcta tgcctcaagg ctctcttaac ttcactgtct cctcttcgga
     2641 taatcctatc cttctcttcc tataaatacc tctccactct tcctcttcct ccaccactac
     2701 aaccaccgca acaaccacca aaaaccctct caaagaaatt tctttttttt cttactttct
     2761 tggtttgtca aataacagta ttcagtcgac tggtaccaac aggctccatg gcttctaatt
     2821 ttactcaatt tgttttggtt gataatggag gaactggaga tgttactgtt gctccttcta
     2881 attttgctaa tggagttgct gaatggattt cttctaattc aagatctcag gcttataagg
     2941 ttacttgttc tgttagacaa tcttctgctc aaaagagaaa gtatactatt aaggttgaag
     3001 ttcctaaggt tgctactcaa actgttggag gagttgaatt gcctgttgct gcttggagat
     3061 cttatttgaa tatggaattg actattccta tttttgctac taattctgat tgtgaattga
     3121 ttgttaaggc tatgcaagga ttgttgaagg atggaaatcc tattccttct gctattgctg
     3181 ctaattctgg aatttattct gctggaggag gaggatctgg aggaggagga tctggaggag
     3241 gaggatctgg acctaagaag aagagaaagg ttgctgctgc tggatcttca ggtggaagag
     3301 ctgatgcttt ggatgatttt gatttggata tgttgggatc tgatgctttg gatgattttg
     3361 atttggatat gttgggatct gatgctttgg atgattttga tttggatatg ttgggatctg
     3421 atgctttgga tgattttgat ttggatatgt tgattaatta atagctgcaa agctttcgtt
     3481 cgtatcatcg gtttcgacaa cgttcgtcaa gttcaatgca tcagtttcat tgcgcacaca
     3541 ccagaatcct actgagtttg agtattatgg cattgggaaa actgtttttc ttgtaccatt
     3601 tgttgtgctt gtaatttact gtgtttttta ttcggttttc gctatcgaac tgtgaaatgg
     3661 aaatggatgg agaagagtta atgaatgata tggtcctttt gttcattctc aaattaatat
     3721 tatttgtttt ttctcttatt tgttgtgtgt tgaatttgaa attataagag atatgcaaac
     3781 attttgtttt gagtaaaaat gtgtcaaatc gtggcctcta atgaccgaag ttaatatgag
     3841 gagtaaaaca cttgtagttg taccattatg cttattcact aggcaacaaa tatattttca
     3901 gacctagaaa agctgcaaat gttactgaat acaagtatgt cctcttgtgt tttagacatt
     3961 tatgaacttt cctttatgta attttccaga atccttgtca gattctaatc attgctttat
     4021 aattatagtt atactcatgg atttgtagtt gagtatgaaa atatttttta atgcatttta
     4081 tgacttgcca attgattgac aacatgcatc aatcgactat gagtaggcaa gatgttctgt
     4141 aggaccttga gaccgaaaag gtggtctcag tattctagac caagcatgca cagctggcgt
     4201 tctcagccag atccccttga caggatatat tggcgggtaa actaagtcgc tgtatgtgtt
     4261 tgtttgagat ctcatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     4321 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     4381 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     4441 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     4501 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     4561 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     4621 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     4681 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     4741 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     4801 gaagccagtt accttcggaa gaagagttgg tagctcttga tccggcaaac aaaccaccgc
     4861 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     4921 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     4981 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     5041 atgaagtttt aaatcaatct aaagtatata tgtgtaacat tggtctagtg attatttgcc
     5101 gactaccttg gtgatctcgc ctttcacgta gtgaacaaat tcttccaact gatctgcgcg
     5161 cgaggccaag cgatcttctt gtccaagata agcctgccta gcttcaagta tgacgggctg
     5221 atactgggcc ggcaggcgct ccattgccca gtcggcagcg acatccttcg gcgcgatttt
     5281 gccggttact gcgctgtacc aaatgcggga caacgtaagc actacatttc gctcatcgcc
     5341 agcccagtcg ggcggcgagt tccatagcgt taaggtttca tttagcgcct caaatagatc
     5401 ctgttcagga accggatcaa agagttcctc cgccgctgga cctaccaagg caacgctatg
     5461 ttctcttgct tttgtcagca agatagccag atcaatgtcg atcgtggctg gctcgaagat
     5521 acctgcaaga atgtcattgc gctgccattc tccaaattgc agttcgcgct tagctggata
     5581 acgccacgga atgatgtcgt cgtgcacaac aatggtgact tctacagcgc ggagaatctc
     5641 gctctctcca ggggaagccg aagtttccaa aaggtcgttg atcaaagctc gccgcgttgt
     5701 ttcatcaagc cttacggtca ccgtaaccag caaatcaata tcactgtgtg gcttcaggcc
     5761 gccatccact gcggagccgt acaaatgtac ggccagcaac gtcggttcga gatggcgctc
     5821 gatgacgcca actacctctg atagttgagt cgatacttcg gcgatcaccg cttccctcat
     5881 aacacccctt gtattactgt ttatgtaagc agacagtttt attgttcatg atgatatatt
     5941 tttatcttgt gcaatgtaac atcagagatt ttgagacaca acgtggcttt gttgaataaa
     6001 tcgaactttt gctgagttga aggatcagat cacgcatctt cccgacaacg cagaccgttc
     6061 cgtggcaaag caaaagttca aaatcaccaa ctggtccacc tacaacaaag ctctcatcaa
     6121 ccgtggctcc ctcactttct ggctggatga tggggcgatt caggcgatcc ccatccaaca
     6181 gcccgccgtc gagcgggct
//
