LOCUS       sequence-68095-e        6956 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Binary vector for Arabidopsis transformation with SapI Golden Gate 
            sites and mCherry seed selection.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6956)
  AUTHORS   Emami S, Yee MC, Dinneny JR
  TITLE     A robust family of Golden Gate Agrobacterium vectors for plant 
            synthetic biology.
  JOURNAL   Front Plant Sci. 2013 Sep 2;4:339. doi: 10.3389/fpls.2013.00339. 
            eCollection 2013.
  PUBMED    24032037
REFERENCE   2  (bases 1 to 6956)
  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: "Front Plant
            Sci."; date: "2013-09-2"; volume: "4:339. doi"; pages: " 
            10.3389/fpls.2013.00339. eCollection 2013"
COMMENT     Sequence Label: pGoldenGate-MCY2
FEATURES             Location/Qualifiers
     source          1..6956
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    3..25
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA 
                     (truncated)"
     promoter        424..769
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     712..732
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     CDS             1677..2387
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEG
                     TQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNF
                     EDGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALK
                     GEIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERA
                     EGRHSTGGMEELYR"
     primer_bind     complement(1822..1840)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2090..2109
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     terminator      2406..2658
                     /label=NOS terminator
                     /note="nopaline synthase terminator and poly(A) signal"
     CDS             complement(2756..3079)
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSLHACRSTLEDPRVPSSNSLAVVLQRRDWENPGVTQLNR
                     LAAHPPFASWRNSEEARTDRPSQQLRSLNGEWRLMRYFLLTHLCGISHRIWCTLSTICS
                     DAA"
     primer_bind     2761..2780
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2974..2996
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     2988..3005
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     2989..3005
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    3006..3062
                     /label=MCS
                     /note="pUC18/19 multiple cloning site"
     primer_bind     complement(3075..3091)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3075..3091)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3088..3110)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3099..3115
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(3123..3129,3130..3147,3148..3153))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3168..3189
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3306..3323)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     polyA_signal    3425..3601
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     primer_bind     complement(3665..3685)
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        complement(3665..3683)
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_feature    3762..3786
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     rep_origin      complement(3877..4465)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3957..3976)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4556..5344)
                     /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      6396..6831
                     /label=pSa ori
                     /note="origin of replication from bacterial plasmid pSa"
ORIGIN
        1 cttggcagga tatattgtgg tgtaacgttg gcgcgcctac tccaaaaatg tcaaagatac
       61 agtctcagaa gaccaaaggg ctattgagac ttttcaacaa agggtaattt cgggaaacct
      121 cctcggattc cattgcccag ctatctgtca cttcatcgaa aggacagtag aaaaggaagg
      181 tggctcctac aaatgccatc attgcgataa aggaaaggct atcattcaag atgcctctgc
      241 cgacagtggt cccaaagatg gacccccacc cacgaggagc atcgtggaaa aagaagacgt
      301 tccaaccacg tcttcaaagc aagtggattg atgtgacatc tccactgacg taagggatga
      361 cgcacaatcc cacccctact ccaaaaatgt caaagataca gtctcagaag accaaagggc
      421 tattgagact tttcaacaaa gggtaatttc gggaaacctc ctcggattcc attgcccagc
      481 tatctgtcac ttcatcgaaa ggacagtaga aaaggaaggt ggctcctaca aatgccatca
      541 ttgcgataaa ggaaaggcta tcattcaaga tgcctctgcc gacagtggtc ccaaagatgg
      601 acccccaccc acgaggagca tcgtggaaaa agaagacgtt ccaaccacgt cttcaaagca
      661 agtggattga tgtgacatct ccactgacgt aagggatgac gcacaatccc actatccttc
      721 gcaagaccct tcctctatat aaggaagttc atttcatttg gagaggacag cccagatcaa
      781 ctagtcttaa gtccggagct agctctagaa tggcaaagga tgtggaagcc gttcccggag
      841 aaggatttca gacaagagac tatcaagatc cgccaccagc tccgtttatt gatggagcgg
      901 agctaaagaa gtggtctttc tacagagcag ttatcgcaga gttcgtagcc actctcctct
      961 tcttatacat caccgttttg acagtcatcg gttacaagat tcagtccgat actgatgccg
     1021 gtggcgtaga ttgcggcgga gttggaatcc tcggtatcgc ttgggccttt ggtggtatga
     1081 tcttcatcct cgtctactgc accgccggta tctctggtgg tcacattaac ccagcggtga
     1141 catttgggct attcttggca cgtaaagtgt cgttacctag ggccctattg tacataatcg
     1201 ctcagtgttt gggtgcgatt tgtggagttg gttttgtcaa agccttccaa agctcttact
     1261 acacccgtta cggaggtgga gccaactctc tagccgatgg ctacagcaca gggaccggtc
     1321 tagccgcaga gatcattggt actttcgttc ttgtctacac cgtcttctct gccactgacc
     1381 ccaaacgtag tgccagagac tcccacgttc cggtgttggc gccacttcca atcggatttg
     1441 ccgtgttcat ggtacatttg gctaccattc ccattaccgg aaccggaatt aacccggcaa
     1501 ggagtttcgg agctgccgta atctacaaca agagcaagcc atgggatgac cactggatat
     1561 tttgggttgg accattcatt ggagctgcga tagctgcatt ctaccaccaa ttcgttctaa
     1621 gagcttcagg ttctaagtct cttggatcat tcagaagtgc tgccaacgtc ggatccatgg
     1681 tgagcaaggg cgaggaggat aacatggcca tcatcaagga gttcatgcgc ttcaaggtgc
     1741 acatggaggg ctccgtgaac ggccacgagt tcgagatcga gggcgagggc gagggccgcc
     1801 cctacgaggg cacccagacc gccaagctga aggtgaccaa gggtggcccc ctgcccttcg
     1861 cctgggacat cctgtcccct cagttcatgt acggctccaa ggcctacgtg aagcaccccg
     1921 ccgacatccc cgactacttg aagctgtcct tccccgaggg cttcaagtgg gagcgcgtga
     1981 tgaacttcga ggacggcggc gtggtgaccg tgacccagga ctcctccctg caggacggcg
     2041 agttcatcta caaggtgaag ctgcgcggca ccaacttccc ctccgacggc cccgtaatgc
     2101 agaagaagac catgggctgg gaggcctcct ccgagcggat gtaccccgag gacggcgccc
     2161 tgaagggcga gatcaagcag aggctgaagc tgaaggacgg cggccactac gacgctgagg
     2221 tcaagaccac ctacaaggcc aagaagcccg tgcagctgcc cggcgcctac aacgtcaaca
     2281 tcaagttgga catcacctcc cacaacgagg actacaccat cgtggaacag tacgaacgcg
     2341 ccgagggccg ccactccacc ggcggcatgg aggagctgta caggtaaagc ggccgcccgg
     2401 ctgcagatcg ttcaaacatt tggcaataaa gtttcttaag attgaatcct gttgccggtc
     2461 ttgcgatgat tatcatataa tttctgttga attacgttaa gcatgtaata attaacatgt
     2521 aatgcatgac gttatttatg agatgggttt ttatgattag agtcccgcaa ttatacattt
     2581 aatacgcgat agaaaacaaa atatagcgcg caaactagga taaattatcg cgcgcggtgt
     2641 catctatgtt actagatcgg ggcgcgccct cgagagtaga agagccggat gccgggagca
     2701 gacaagcccg tcagggcgcg tcagcgggtg ttggcgggtg tcggggctgg cttaactatg
     2761 cggcatcaga gcagattgta ctgagagtgc accatatgcg gtgtgaaata ccgcacagat
     2821 gcgtaaggag aaaataccgc atcaggcgcc attcgccatt caggctgcgc aactgttggg
     2881 aagggcgatc ggtgcgggcc tcttcgctat tacgccagct ggcgaaaggg ggatgtgctg
     2941 caaggcgatt aagttgggta acgccagggt tttcccagtc acgacgttgt aaaacgacgg
     3001 ccagtgaatt cgagctcggt acccggggat cctctagagt cgacctgcag gcatgcaggc
     3061 ttggcgtaat catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca
     3121 cacaacatac gagccggaag cataaagtgt aaagcctggg gtgcctaatg agtgagctaa
     3181 ctcacattaa ttgcgttgcg ctcactgccc gctttccagt cgggaaacct gtcgtgccag
     3241 ctgcattaat gaatcggcca acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc
     3301 gcttcctcgc tcactgactc gctgcgctcg gtcgttcggc tgcggcgagc ggtatcgctc
     3361 ttcatggaag cttgtcggta cgctgaaatc accagtctct ctctacaaat ctatctctct
     3421 ctattttctc cataaataat gtgtgagtag tttcccgata agggaaatta gggttcttat
     3481 agggtttcgc tcatgtgttg agcatataag aaacccttag tatgtatttg tatttgtaaa
     3541 atacttctat caataaaatt tctaattcct aaaaccaaaa tccagtacta aaatccagat
     3601 cgatgggcta gagcggacta gttctagagc ggccgccacc gcggtggagc tccagctttt
     3661 gttcccttta gtgagggtta attgcgcgct agctcactca aaggcggtaa tacggttatc
     3721 cacagaatca ggggataacg caggaaagaa catgaaggcc ttgacaggat atattggcgg
     3781 gtaaactaag tcgctgtatg tgtttgtttg agatctcatg tgagcaaaag gccagcaaaa
     3841 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     3901 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     3961 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     4021 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     4081 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     4141 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     4201 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     4261 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     4321 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaagaagag ttggtagctc
     4381 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     4441 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     4501 tcagtggaac gaaaactcac gttaagggat tttggtcatg aattcttgtt agacattatt
     4561 tgccgactac cttggtgatc tcgcctttca cgtagtgaac aaattcttcc aactgatctg
     4621 cgcgcgaggc caagcgatct tcttgtccaa gataagcctg cctagcttca agtatgacgg
     4681 gctgatactg ggccggcagg cgctccattg cccagtcggc agcgacatcc ttcggcgcga
     4741 ttttgccggt tactgcgctg taccaaatgc gggacaacgt aagcactaca tttcgctcat
     4801 cgccagccca gtcgggcggc gagttccata gcgttaaggt ttcatttagc gcctcaaata
     4861 gatcctgttc aggaaccgga tcaaagagtt cctccgccgc tggacctacc aaggcaacgc
     4921 tatgttctct tgcttttgtc agcaagatag ccagatcaat gtcgatcgtg gctggctcga
     4981 agatacctgc aagaatgtca ttgcgctgcc attctccaaa ttgcagttcg cgcttagctg
     5041 gataacgcca cggaatgatg tcgtcgtgca caacaatggt gacttctaca gcgcggagaa
     5101 tctcgctctc tccaggggaa gccgaagttt ccaaaaggtc gttgatcaaa gctcgccgcg
     5161 ttgtttcatc aagccttacg gtcaccgtaa ccagcaaatc aatatcactg tgtggcttca
     5221 ggccgccatc cactgcggag ccgtacaaat gtacggccag caacgtcggt tcgagatggc
     5281 gctcgatgac gccaactacc tctgatagtt gagtcgatac ttcggcgatc accgcttccc
     5341 tcatgatgtt taactcctga attaagccgc gccgcgaagc ggtgtggctt gaatgaattg
     5401 ttaggcgtca tcctgtgctc ccgagaacca gtaccagtac atcgctgttt cgttcgagac
     5461 ttgaggtcta gttttatacg tgaacaggtc aatgccgccg agagtaaagc cacatttgcg
     5521 tacaaattgc aggcaggtac attgttcgtt tgtgtctcta atcgtatgcc aaggagctgt
     5581 ctgcttagtg cccacttttt cgcaaaattc gatgagactg tgcgcgactc ctttgcctcg
     5641 gtgcgtgtgc gacacaacaa tgtgttcgat agaggctaga tcgttcccat gttgagttga
     5701 gttcaatctt cccgacaagc tcttggtcga tgaatgtgcc atagcaagca gagtcttcat
     5761 cagagtcatc atccgagatg taatccttcc ggtaggggct cacacttctg gtagatagtt
     5821 caaagccttg gtcggatagg tgcacatcga acacttcacg aacaatgaaa tggttctcag
     5881 catccaatgt ttccgccacc tgctcaggga tcaccgaaat cttcatatga cgcctaacgc
     5941 ctggcacagc ggatcgcaaa cctggcgcgg cttttggcac aaaaggcgtg acaggtttgc
     6001 gaatccgttg ctgccacttg tttaatagac tggatggagg cggataaagt tgcaggacca
     6061 cttctgcgct cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag
     6121 cgtggatctc gcggtatcat tgcagcactg ccatgatgat atatttttat cttgtgcaat
     6181 gtaacatcag agattttgag acacaacgtg gctttgttga ataaatcgaa cttttgctga
     6241 gttgaaggat cagatcacgc atcttcccga caacgcagac cgttccgtgg caaagcaaaa
     6301 gttcaaaatc accaactggt ccacctacaa caaagctctc atcaaccgtg gctccctcac
     6361 tttctggctg gatgatgggg cgattcaggc gatccccatc caacagcccg ccgtcgagcg
     6421 ggctttttta tccccggaag cctgtggata gagggtagtt atccacgtga aaccgctaat
     6481 gccccgcaaa gccttgattc acggggcttt ccggcccgct ccaaaaacta tccacgtgaa
     6541 atcgctaatc agggtacgtg aaatcgctaa tcggagtacg tgaaatcgct aataaggtca
     6601 cgtgaaatcg ctaatcaaaa aggcacgtga gaacgctaat agccctttca gatcaacagc
     6661 ttgcaaacac ccctcgctcc ggcaagtagt tacagcaagt agtatgttca attagctttt
     6721 caattatgaa tatatatatc aattattggt cgcccttggc ttgtggacaa tgcgctacgc
     6781 gcaccggctc cgcccgtgga caaccgcaag cggttgccca ccgtcgagcg cctttgccca
     6841 caacccggcg gccggccgca acagatcgtt ttataaattt ttttttttga aaaagaaaaa
     6901 gcccgaaagg cggcaacctc tcgggcttct ggatttccga tccccggaat tagact
//
