LOCUS       sequence-239401-        4933 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Binary expression vector containing 35S: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 4933)
  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 4933)
  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: pMH204
FEATURES             Location/Qualifiers
     source          1..4933
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        61..406
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     349..369
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     CDS             457..843
                     /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             898..918
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             949..1098
                     /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      1117..1758
                     /label=E9 terminator
                     /note="terminator and polyadenylation signal from the pea 
                     rbcS-E9 gene"
     misc_feature    1864..1888
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     rep_origin      complement(1979..2567)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2059..2078)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(2738..3526)
                     /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      3817..4252
                     /label=pSa ori
                     /note="origin of replication from bacterial plasmid pSa"
     misc_feature    4379..4401
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA 
                     (truncated)"
ORIGIN
        1 cgacacactt gtctactcca aaaatatcaa agatacagtc tcagaagacc aaagggcaat
       61 tgagactttt caacaaaggg taatatccgg aaacctcctc ggattccatt gcccagctat
      121 ctgtcacttt attgtgaaga tagtggaaaa ggaaggtggc tcctacaaat gccatcattg
      181 cgataaagga aaggccatcg ttgaagatgc ctctgccgac agtggtccca aagatggacc
      241 cccacccacg aggagcatcg tggaaaaaga agacgttcca accacgtctt caaagcaagt
      301 ggattgatgt gatatctcca ctgacgtaag ggatgacgca caatcccact atccttcgca
      361 agacccttcc tctatataag gaagttcatt tcatttggag agaacacggg ggacgagcta
      421 acagtattca gtcgactggt accaacaggc tccatggctt ctaattttac tcaatttgtt
      481 ttggttgata atggaggaac tggagatgtt actgttgctc cttctaattt tgctaatgga
      541 gttgctgaat ggatttcttc taattcaaga tctcaggctt ataaggttac ttgttctgtt
      601 agacaatctt ctgctcaaaa gagaaagtat actattaagg ttgaagttcc taaggttgct
      661 actcaaactg ttggaggagt tgaattgcct gttgctgctt ggagatctta tttgaatatg
      721 gaattgacta ttcctatttt tgctactaat tctgattgtg aattgattgt taaggctatg
      781 caaggattgt tgaaggatgg aaatcctatt ccttctgcta ttgctgctaa ttctggaatt
      841 tattctgctg gaggaggagg atctggagga ggaggatctg gaggaggagg atctggacct
      901 aagaagaaga gaaaggttgc tgctgctgga tcttcaggtg gaagagctga tgctttggat
      961 gattttgatt tggatatgtt gggatctgat gctttggatg attttgattt ggatatgttg
     1021 ggatctgatg ctttggatga ttttgatttg gatatgttgg gatctgatgc tttggatgat
     1081 tttgatttgg atatgttgat taattaatag ctgcaaagct ttcgttcgta tcatcggttt
     1141 cgacaacgtt cgtcaagttc aatgcatcag tttcattgcg cacacaccag aatcctactg
     1201 agtttgagta ttatggcatt gggaaaactg tttttcttgt accatttgtt gtgcttgtaa
     1261 tttactgtgt tttttattcg gttttcgcta tcgaactgtg aaatggaaat ggatggagaa
     1321 gagttaatga atgatatggt ccttttgttc attctcaaat taatattatt tgttttttct
     1381 cttatttgtt gtgtgttgaa tttgaaatta taagagatat gcaaacattt tgttttgagt
     1441 aaaaatgtgt caaatcgtgg cctctaatga ccgaagttaa tatgaggagt aaaacacttg
     1501 tagttgtacc attatgctta ttcactaggc aacaaatata ttttcagacc tagaaaagct
     1561 gcaaatgtta ctgaatacaa gtatgtcctc ttgtgtttta gacatttatg aactttcctt
     1621 tatgtaattt tccagaatcc ttgtcagatt ctaatcattg ctttataatt atagttatac
     1681 tcatggattt gtagttgagt atgaaaatat tttttaatgc attttatgac ttgccaattg
     1741 attgacaaca tgcatcaatc gactatgagt aggcaagatg ttctgtagga ccttgagacc
     1801 gaaaaggtgg tctcagtatt ctagaccaag catgcacagc tggcgttctc agccagatcc
     1861 ccttgacagg atatattggc gggtaaacta agtcgctgta tgtgtttgtt tgagatctca
     1921 tgtgagcaaa aggccagcaa aaggccagga accgtaaaaa ggccgcgttg ctggcgtttt
     1981 tccataggct ccgcccccct gacgagcatc acaaaaatcg acgctcaagt cagaggtggc
     2041 gaaacccgac aggactataa agataccagg cgtttccccc tggaagctcc ctcgtgcgct
     2101 ctcctgttcc gaccctgccg cttaccggat acctgtccgc ctttctccct tcgggaagcg
     2161 tggcgctttc tcatagctca cgctgtaggt atctcagttc ggtgtaggtc gttcgctcca
     2221 agctgggctg tgtgcacgaa ccccccgttc agcccgaccg ctgcgcctta tccggtaact
     2281 atcgtcttga gtccaacccg gtaagacacg acttatcgcc actggcagca gccactggta
     2341 acaggattag cagagcgagg tatgtaggcg gtgctacaga gttcttgaag tggtggccta
     2401 actacggcta cactagaaga acagtatttg gtatctgcgc tctgctgaag ccagttacct
     2461 tcggaagaag agttggtagc tcttgatccg gcaaacaaac caccgctggt agcggtggtt
     2521 tttttgtttg caagcagcag attacgcgca gaaaaaaagg atctcaagaa gatcctttga
     2581 tcttttctac ggggtctgac gctcagtgga acgaaaactc acgttaaggg attttggtca
     2641 tgagattatc aaaaaggatc ttcacctaga tccttttaaa ttaaaaatga agttttaaat
     2701 caatctaaag tatatatgtg taacattggt ctagtgatta tttgccgact accttggtga
     2761 tctcgccttt cacgtagtga acaaattctt ccaactgatc tgcgcgcgag gccaagcgat
     2821 cttcttgtcc aagataagcc tgcctagctt caagtatgac gggctgatac tgggccggca
     2881 ggcgctccat tgcccagtcg gcagcgacat ccttcggcgc gattttgccg gttactgcgc
     2941 tgtaccaaat gcgggacaac gtaagcacta catttcgctc atcgccagcc cagtcgggcg
     3001 gcgagttcca tagcgttaag gtttcattta gcgcctcaaa tagatcctgt tcaggaaccg
     3061 gatcaaagag ttcctccgcc gctggaccta ccaaggcaac gctatgttct cttgcttttg
     3121 tcagcaagat agccagatca atgtcgatcg tggctggctc gaagatacct gcaagaatgt
     3181 cattgcgctg ccattctcca aattgcagtt cgcgcttagc tggataacgc cacggaatga
     3241 tgtcgtcgtg cacaacaatg gtgacttcta cagcgcggag aatctcgctc tctccagggg
     3301 aagccgaagt ttccaaaagg tcgttgatca aagctcgccg cgttgtttca tcaagcctta
     3361 cggtcaccgt aaccagcaaa tcaatatcac tgtgtggctt caggccgcca tccactgcgg
     3421 agccgtacaa atgtacggcc agcaacgtcg gttcgagatg gcgctcgatg acgccaacta
     3481 cctctgatag ttgagtcgat acttcggcga tcaccgcttc cctcataaca ccccttgtat
     3541 tactgtttat gtaagcagac agttttattg ttcatgatga tatattttta tcttgtgcaa
     3601 tgtaacatca gagattttga gacacaacgt ggctttgttg aataaatcga acttttgctg
     3661 agttgaagga tcagatcacg catcttcccg acaacgcaga ccgttccgtg gcaaagcaaa
     3721 agttcaaaat caccaactgg tccacctaca acaaagctct catcaaccgt ggctccctca
     3781 ctttctggct ggatgatggg gcgattcagg cgatccccat ccaacagccc gccgtcgagc
     3841 gggctttttt atccccggaa gcctgtggat agagggtagt tatccacgtg aaaccgctaa
     3901 tgccccgcaa agccttgatt cacggggctt tccggcccgc tccaaaaact atccacgtga
     3961 aatcgctaat cagggtacgt gaaatcgcta atcggagtac gtgaaatcgc taataaggtc
     4021 acgtgaaatc gctaatcaaa aaggcacgtg agaacgctaa tagccctttc agatcaacag
     4081 cttgcaaaca cccctcgctc cggcaagtag ttacagcaag tagtatgttc aattagcttt
     4141 tcaattatga atatatatat caattattgg tcgcccttgg cttgtggaca atgcgctacg
     4201 cgcaccggct ccgcccgtgg acaaccgcaa gcggttgccc accgtcgagc gcctttgccc
     4261 acaacccggc ggccggccgc aacagatcgt tttataaatt tttttttttg aaaaagaaaa
     4321 agcccgaaag gcggcaacct ctcgggcttc tggatttccg atccccggaa ttagatcttg
     4381 gcaggatata ttgtggtgta acgttctcga actaggctcg gacgaagtaa gcggccgcaa
     4441 ccttaattaa acacggcgcg ccaacagcga tcgcaccagg taccacctcc gatccattcg
     4501 gtccccagat tagccttttc aatttcagaa agaatgctaa cccacagatg gttagagagg
     4561 cttacgcagc acgtctcatc aagacgatct acccgagcaa taatctccag gaaatcaaat
     4621 accttcccaa gaaggttaaa gatgcagtca aaagattcag gactaactgc atcaagaaca
     4681 cagagaaaga tatatttctc aagatcagaa gtactattcc agtatggacg attcaaggct
     4741 tgcttcacaa accaaggcaa gtaatagaga ttggagtctc taaaaaggta gttcccactg
     4801 aatcaaaggc catggagtca aagattcaaa tagaggacct aacagaactc gccgtaaaga
     4861 ctggcgaaca gttcatacag agtctcttac gactcaatga caagaagaaa atcttcgtca
     4921 acatggtgga gca
//
