LOCUS       sequence-182146-        5608 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Transformation, Type: non-T-DNA, Plant Selection: 2x35S:npt II, 
            Viral Replication: none.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5608)
  AUTHORS   Cermak T, Curtin SJ, Gil-Humanes J, Cegan R, Kono TJY, Konecna E, 
            Belanto JJ, Starker CG, Mathre JW, Greenstein RL, Voytas DF
  TITLE     A multi-purpose toolkit to enable advanced genome engineering in 
            plants.
  JOURNAL   Plant Cell. 2017 May 18. pii: tpc.00922.2016. doi: 
            10.1105/tpc.16.00922.
  PUBMED    28522548
REFERENCE   2  (bases 1 to 5608)
  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: "Plant Cell.
            2017 May 18. pii: tpc.00922.2016. doi: 10.1105/tpc.16.00922."
COMMENT     Sequence Label: pTRANS_120
FEATURES             Location/Qualifiers
     source          1..5608
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      84..672
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     573..592
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     826..843
                     /label=L4440
                     /note="L4440 vector, forward primer"
     polyA_signal    872..1046
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     CDS             complement(1103..1900)
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MGIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGR
                     PVLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLL
                     SSHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEH
                     QGLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDI
                     ALATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(1214..1233)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     1824..1843
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(1963..2639)
                     /label=CaMV 35S promoter (enhanced)
                     /note="cauliflower mosaic virus 35S promoter with a 
                     duplicated enhancer region"
     primer_bind     complement(2006..2026)
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     protein_bind    2736..2860
                     /gene="mutant version of attR"
                     /label=attR1
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     promoter        join(2885..2890,2891..2908,2909..2915)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     CDS             2969..3649
                     /codon_start=1
                     /product="chloramphenicol acetyltransferase"
                     /label=CmR
                     /note="confers resistance to chloramphenicol"
                     /translation="MEKKITGYTTVDISQWHRKEHFEAFQSVAQCTYNQTVQLDITAFL
                     KTVKKNKHKFYPAFIHILARLMNAHPEFRMAMKDGELVIWDSVHPCYTVFHEQTETFSS
                     LWSEYHDDFRQFLHIYSQDVACYGENLAYFPKGFIENMFFVSANPWVSFTSFDLNVANM
                     DNFFAPVFTMGKYYTQGDKVLMPLAIQVHHAVCDGFHVGRMLNELQQYCDEWQAGRNLE
                     DPAY"
     CDS             3969..4274
                     /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
                     VPRELYPVVHIGDESWRMMTTDMASVPASVIGEEVADLSHRENDIKNAINLMFWGI"
     primer_bind     4153..4172
                     /label=ccdB-fwd
                     /note="ccdB gene, forward primer"
     protein_bind    complement(4315..4439)
                     /gene="mutant version of attR"
                     /label=attR2
                     /bound_moiety="LR Clonase(TM)"
                     /note="recombination site for the Gateway(R) LR reaction"
     CDS             4808..5599
                     /codon_start=1
                     /gene="aadA"
                     /product="aminoglycoside adenylyltransferase (Murphy, 
                     1985)"
                     /label=SmR
                     /note="confers resistance to spectinomycin and 
                     streptomycin"
                     /translation="MREAVIAEVSTQLSEVVGVIERHLEPTLLAVHLYGSAVDGGLKPH
                     SDIDLLVTVTVRLDETTRRALINDLLETSASPGESEILRAVEVTIVVHDDIIPWRYPAK
                     RELQFGEWQRNDILAGIFEPATIDIDLAILLTKAREHSVALVGPAAEELFDPVPEQDLF
                     EALNETLTLWNSPPDWAGDERNVVLTLSRIWYSAVTGKIAPKDVAADWAMERLPAQYQP
                     VILEARQAYLGQEEDRLASRADQLEEFVHYVKGEITKVVGK"
ORIGIN
        1 cacccatgac caaaatccct taacgtgagt tacgcgtcgt tccactgagc gtcagacccc
       61 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
      121 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
      181 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
      241 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
      301 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
      361 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
      421 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
      481 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
      541 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
      601 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
      661 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
      721 tttgctcaca tgttctttcc tgcgttatcc cctgattctg tggataaccg tattaccgcc
      781 tttgagtgag ctgataccgc tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc
      841 gaggaagcgg gagagcgccc ataattcggg ggatctggat tttagtactg gattttggtt
      901 ttaggaatta gaaattttat tgatagaagt attttacaaa tacaaataca tactaagggt
      961 ttcttatatg ctcaacacat gagcgaaacc ctataggaac cctaattccc ttatctggga
     1021 actactcaca cattattatg gagaaactcg agcttgtcga tcgactctag ctagaggatc
     1081 gatccgaacc ccagagtccc gctcagaaga actcgtcaag aaggcgatag aaggcgatgc
     1141 gctgcgaatc gggagcggcg ataccgtaaa gcacgaggaa gcggtcagcc cattcgccgc
     1201 caagttcttc agcaatatca cgggtagcca acgctatgtc ctgatagcgg tccgccacac
     1261 ccagccggcc acagtcgatg aatccagaaa agcggccatt ttccaccatg atattcggca
     1321 agcaggcatc gccatgtgtc acgacgagat cctcgccgtc gggcatgcgc gccttgagcc
     1381 tggcgaacag ttcggctggc gcgagcccct gatgttcttc gtccagatca tcctgatcga
     1441 caagaccggc ttccatccga gtacgtgctc gctcgatgcg atgtttcgct tggtggtcga
     1501 atgggcaggt agccggatca agcgtatgca gccgccgcat tgcatcagcc atgatggata
     1561 ctttctcggc aggagcaagg tgagatgaca ggagatcctg ccccggcact tcgcccaata
     1621 gcagccagtc ccttcccgct tcagtgacaa cgtcgagcac agctgcgcaa ggaacgcccg
     1681 tcgtggccag ccacgatagc cgcgctgcct cgtcctggag ttcattcagg gcaccggaca
     1741 ggtcggtctt gacaaaaaga accgggcgcc cctgcgctga cagccgaaac acggcggcat
     1801 cagagcagcc gattgtctgt tgtgcccagt catagccgaa tagcctctcc acccaagcgg
     1861 ccggagaacc tgcgtgcaat ccatcttgtt caatccccat ggtcgatcga cagatctgcg
     1921 aaagctcgag agagatagat ttgtagagag agactggtga tttcagcgtg tcctctccaa
     1981 atgaaatgaa cttccttata tagaggaagg gtcttgcgaa ggatagtggg attgtgcgtc
     2041 atcccttacg tcagtggaga tatcacatca atccacttgc tttgaagacg tggttggaac
     2101 gtcttctttt tccacgatgc tcctcgtggg tgggggtcca tctttgggac cactgtcggc
     2161 agaggcatct tgaacgatag cctttccttt atcgcaatga tggcatttgt aggtgccacc
     2221 ttccttttct actgtccttt tgatgaagtg acagatagct gggcaatgga atccgaggag
     2281 gtttcccgat attacccttt gttgaaaagt ctcaatagcc ctttggtctt ctgagactgt
     2341 atctttgata ttcttggagt agacgagagt gtcgtgctcc accatgttca catcaatcca
     2401 cttgctttga agacgtggtt ggaacgtctt ctttttccac gatgctcctc gtgggtgggg
     2461 gtccatcttt gggaccactg tcggcagagg catcttgaac gatagccttt cctttatcgc
     2521 aatgatggca tttgtaggtg ccaccttcct tttctactgt ccttttgatg aagtgacaga
     2581 tagctgggca atggaatccg aggaggtttc ccgatattac cctttgttga aaagtctcaa
     2641 tagccctttg gtcttctgag actgtatctt tgatattctt ggagtagacg agagtgtcgt
     2701 gctccaccat gcggcgcgcc gatcaaaagc aggtgacaag tttgtacaaa aaagctgaac
     2761 gagaaacgta aaatgatata aatatcaata tattaaatta gattttgcat aaaaaacaga
     2821 ctacataata ctgtaaaaca caacatatcc agtcactatg gcggccgcat taggcacccc
     2881 aggctttaca ctttatgctt ccggctcgta taatgtgtgg attttgagtt aggatccgtc
     2941 gagattttca ggagctaagg aagctaaaat ggagaaaaaa atcactggat ataccaccgt
     3001 tgatatatcc caatggcatc gtaaagaaca ttttgaggca tttcagtcag tcgctcaatg
     3061 tacctataac cagaccgttc agctggatat tacggccttt ttaaagaccg taaagaaaaa
     3121 taagcacaag ttttatccgg cctttattca cattcttgcc cgcctgatga atgctcatcc
     3181 ggaattccgt atggcaatga aagacggtga gctggtgata tgggatagtg ttcacccttg
     3241 ttacaccgtt ttccatgagc aaactgaaac gttttcatcg ctctggagtg aataccacga
     3301 cgatttccgg cagtttctac acatatattc gcaagatgtg gcgtgttacg gtgaaaacct
     3361 ggcctatttc cctaaagggt ttattgagaa tatgtttttc gtctcagcca atccctgggt
     3421 gagtttcacc agttttgatt taaacgtggc caatatggac aacttcttcg cccccgtttt
     3481 caccatgggc aaatattata cgcaaggcga caaggtgctg atgccgctgg cgattcaggt
     3541 tcatcatgcc gtttgtgatg gcttccatgt cggcagaatg cttaatgaat tacaacagta
     3601 ctgcgatgag tggcaggcgg ggcgtaatct agaggatccg gcttactaaa agccagataa
     3661 cagtatgcgt atttgcgcgc tgatttttgc ggtataagaa tatatactga tatgtatacc
     3721 agaagtatgt caaaaagagg tatgctatga agcagcgtat tacagtgaca gttgacagcg
     3781 acagctatca gttgctcaag gcatatatga tgtcaatatc tccggtctgg taagcacaac
     3841 catgcagaat gaagcccgtc gtctgcgtgc cgaacgctgg aaagcggaaa atcaggaagg
     3901 gatggctgag gtcgcccggt ttattgaaat gaacggctct tttgctgacg agaacagggg
     3961 ctggtgaaat gcagtttaag gtttacacct ataaaagaga gagccgttat cgtctgtttg
     4021 tggatgtaca gagtgatatt attgacacgc ccgggcgacg gatggtgatc cccctggcca
     4081 gtgcacgtct gctgtcagat aaagtccccc gtgaacttta cccggtggtg catatcgggg
     4141 atgaaagctg gcgcatgatg accaccgata tggccagtgt gccggcctcc gttatcgggg
     4201 aagaagtggc tgatctcagc caccgcgaaa atgacatcaa aaacgccatt aacctgatgt
     4261 tctggggaat ataaatgtca ggctccctta tacacagcca gtctgcaggt cgaccatagt
     4321 gactggatat gttgtgtttt acagtattat gtagtctgtt ttttatgcaa aatctaattt
     4381 aatatattga tatttatatc attttacgtt tctcgttcag ctttcttgta caaagtggtc
     4441 acctgcaaaa agtgccagcc aggacagaaa tgcctcgact tcgctgctac ccaaggttgc
     4501 cgggtgacgc acaccgtgga aacggatgaa ggcacgaacc cagtggacat aagcctgttc
     4561 ggttcgtaag ctgtaatgca agtagcgtat gcgctcacgc aactggtcca gaaccttgac
     4621 cgaacgcagc ggtggtaacg gcgcagtggc ggttttcatg gcttgttatg actgtttttt
     4681 tggggtacag tctatgcctc gggcatccaa gcagcaagcg cgttacgccg tgggtcgatg
     4741 tttgatgtta tggagcagca acgatgttac gcagcagggc agtcgcccta aaacaaagtt
     4801 aaacattatg agggaagcgg tgatcgccga agtatcgact caactatcag aggtagttgg
     4861 cgtcatcgag cgccatctcg aaccgacgtt gctggccgta catttgtacg gctccgcagt
     4921 ggatggcggc ctgaagccac acagtgatat tgatttgctg gttacggtga ccgtaaggct
     4981 tgatgaaaca acgcggcgag ctttgatcaa cgaccttttg gaaacttcgg cttcccctgg
     5041 agagagcgag attctccgcg ctgtagaagt caccattgtt gtgcacgacg acatcattcc
     5101 gtggcgttat ccagctaagc gcgaactgca atttggagaa tggcagcgca atgacattct
     5161 tgcaggtatc ttcgagccag ccacgatcga cattgatctg gctatcttgc tgacaaaagc
     5221 aagagaacat agcgttgcct tggtaggtcc agcggcggag gaactctttg atccggttcc
     5281 tgaacaggat ctatttgagg cgctaaatga aaccttaacg ctatggaact cgccgcccga
     5341 ctgggctggc gatgagcgaa atgtagtgct tacgttgtcc cgcatttggt acagcgcagt
     5401 aaccggcaaa atcgcgccga aggatgtcgc tgccgactgg gcaatggagc gcctgccggc
     5461 ccagtatcag cccgtcatac ttgaagctag acaggcttat cttggacaag aagaagatcg
     5521 cttggcctcg cgcgcagatc agttggaaga atttgtccac tacgtgaaag gcgagatcac
     5581 caaggtagtc ggcaaataac cctcgagc
//
