LOCUS       sequence-477314-        5535 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5535)
  TITLE     Plasmids to assay for nuclease cleavage in planta
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 5535)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pGreenII 0579-1 (35S::LUC)
FEATURES             Location/Qualifiers
     source          1..5535
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    30..54
                     /label=RB T-DNA repeat
                     /note="right border repeat from nopaline C58 T-DNA"
     primer_bind     159..176
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    293..314
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(329..334,335..352,353..359)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    367..383
                     /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)."
     primer_bind     372..394
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     391..407
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     391..407
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     426..446
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        428..446
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     primer_bind     complement(500..519)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(501..519)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(529..546)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(529..545)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(538..560)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     promoter        763..1108
                     /label=CaMV 35S promoter
                     /note="strong constitutive promoter from cauliflower mosaic
                     virus"
     primer_bind     1051..1071
                     /label=35S promoter
                     /note="CaMV 35S promoter, forward primer"
     CDS             1119..2771
                     /codon_start=1
                     /gene="luc+"
                     /product="firefly luciferase"
                     /label=luciferase
                     /note="enhanced luc+ version of the luciferase gene"
                     /translation="MEDAKNIKKGPAPFYPLEDGTAGEQLHKAMKRYALVPGTIAFTDA
                     HIEVDITYAEYFEMSVRLAEAMKRYGLNTNHRIVVCSENSLQFFMPVLGALFIGVAVAP
                     ANDIYNERELLNSMGISQPTVVFVSKKGLQKILNVQKKLPIIQKIIIMDSKTDYQGFQS
                     MYTFVTSHLPPGFNEYDFVPESFDRDKTIALIMNSSGSTGLPKGVALPHRTACVRFSHA
                     RDPIFGNQIIPDTAILSVVPFHHGFGMFTTLGYLICGFRVVLMYRFEEELFLRSLQDYK
                     IQSALLVPTLFSFFAKSTLIDKYDLSNLHEIASGGAPLSKEVGEAVAKRFHLPGIRQGY
                     GLTETTSAILITPEGDDKPGAVGKVVPFFEAKVVDLDTGKTLGVNQRGELCVRGPMIMS
                     GYVNNPEATNALIDKDGWLHSGDIAYWDEDEHFFIVDRLKSLIKYKGYQVAPAELESIL
                     LQHPNIFDAGVAGLPDDDAGELPAAVVVLEHGKTMTEKEIVDYVASQVTTAKKLRGGVV
                     FVDEVPKGLTGKLDARKIREILIKAKKGGKIAV"
     primer_bind     complement(1185..1203)
                     /label=LucNrev
                     /note="Firefly luciferase, reverse primer"
     polyA_signal    2841..3017
                     /label=CaMV poly(A) signal
                     /note="cauliflower mosaic virus polyadenylation signal"
     misc_feature    3025..3047
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA 
                     (truncated)"
     rep_origin      complement(3114..3702)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3194..3213)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(3873..4688)
                     /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(3873..4688)
                     /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     4596..4615
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     4680..4699
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     rep_origin      4979..5414
                     /label=pSa ori
                     /note="origin of replication from bacterial plasmid pSa"
ORIGIN
        1 gatctcaaac aaacacatac agcgacttag tttacccgcc aatatatcct gtcaaggcct
       61 tcatgttctt tcctgcgtta tcccctgatt ctgtggataa ccgtattacc gcctttgagt
      121 gagctgatac cgctcgccgc agccgaacga ccgagcgcag cgagtcagtg agcgaggaag
      181 cggaagagcg cccaatacgc aaaccgcctc tccccgcgcg ttggccgatt cattaatgca
      241 gctggcacga caggtttccc gactggaaag cgggcagtga gcgcaacgca attaatgtga
      301 gttagctcac tcattaggca ccccaggctt tacactttat gcttccggct cgtatgttgt
      361 gtggaattgt gagcggataa caatttcaca caggaaacag ctatgaccat gattacgcca
      421 agcgcgcaat taaccctcac taaagggaac aaaagctgga gctccaccgc ggtggcggcc
      481 gctctaggta cccaattcgc cctatagtga gtcgtattac gcgcgctcac tggccgtcgt
      541 tttacaacgt cgtgactggg aaaaccctgg cgttacccaa cttaatcgcc ttgcagcaca
      601 tccccctttc gccagctggc gtaatagcga agaggcccgc accgatcgcc cttcccaaca
      661 gttgcgcagc ctgaatggcg aatggaaatt gtaagcgtta atgttatcgt acccctactc
      721 caaaaatgtc aaagatacag tctcagaaga ccaaagggct attgagactt ttcaacaaag
      781 ggtaatttcg ggaaacctcc tcggattcca ttgcccagct atctgtcact tcatcgaaag
      841 gacagtagaa aaggaaggtg gctcctacaa atgccatcat tgcgataaag gaaaggctat
      901 cattcaagat gcctctgccg acagtggtcc caaagatgga cccccaccca cgaggagcat
      961 cgtggaaaaa gaagacgttc caaccacgtc ttcaaagcaa gtggattgat gtgacatctc
     1021 cactgacgta agggatgacg cacaatccca ctatccttcg caagaccctt cctctatata
     1081 aggaagttca tttcatttgg agaggacagc ccaagctcat ggaagacgcc aaaaacataa
     1141 agaaaggccc ggcgccattc tatccgctgg aagatggaac cgctggagag caactgcata
     1201 aggctatgaa gagatacgcc ctggttcctg gaacaattgc ttttacagat gcacatatcg
     1261 aggtggacat cacttacgct gagtacttcg aaatgtccgt tcggttggca gaagctatga
     1321 aacgatatgg gctgaataca aatcacagaa tcgtcgtatg cagtgaaaac tctcttcaat
     1381 tctttatgcc ggtgttgggc gcgttattta tcggagttgc agttgcgccc gcgaacgaca
     1441 tttataatga acgtgaattg ctcaacagta tgggcatttc gcagcctacc gtggtgttcg
     1501 tttccaaaaa ggggttgcaa aaaattttga acgtgcaaaa aaagctccca atcatccaaa
     1561 aaattattat catggattct aaaacggatt accagggatt tcagtcgatg tacacgttcg
     1621 tcacatctca tctacctccc ggttttaatg aatacgattt tgtgccagag tccttcgata
     1681 gggacaagac aattgcactg atcatgaact cctctggatc tactggtctg cctaaaggtg
     1741 tcgctctgcc tcatagaact gcctgcgtga gattctcgca tgccagagat cctatttttg
     1801 gcaatcaaat cattccggat actgcgattt taagtgttgt tccattccat cacggttttg
     1861 gaatgtttac tacactcgga tatttgatat gtggatttcg agtcgtctta atgtatagat
     1921 ttgaagaaga gctgtttctg aggagccttc aggattacaa gattcaaagt gcgctgctgg
     1981 tgccaaccct attctccttc ttcgccaaaa gcactctgat tgacaaatac gatttatcta
     2041 atttacacga aattgcttct ggtggcgctc ccctctctaa ggaagtcggg gaagcggttg
     2101 ccaagaggtt ccatctgcca ggtatcaggc aaggatatgg gctcactgag actacatcag
     2161 ctattctgat tacacccgag ggggatgata aaccgggcgc ggtcggtaaa gttgttccat
     2221 tttttgaagc gaaggttgtg gatctggata ccgggaaaac gctgggcgtt aatcaaagag
     2281 gcgaactgtg tgtgagaggt cctatgatta tgtccggtta tgtaaacaat ccggaagcga
     2341 ccaacgcctt gattgacaag gatggatggc tacattctgg agacatagct tactgggacg
     2401 aagacgaaca cttcttcatc gttgaccgcc tgaagtctct gattaagtac aaaggctatc
     2461 aggtggctcc cgctgaattg gaatccatct tgctccaaca ccccaacatc ttcgacgcag
     2521 gtgtcgcagg tcttcccgac gatgacgccg gtgaacttcc cgccgccgtt gttgttttgg
     2581 agcacggaaa gacgatgacg gaaaaagaga tcgtggatta cgtcgccagt caagtaacaa
     2641 ccgcgaaaaa gttgcgcgga ggagttgtgt ttgtggacga agtaccgaaa ggtcttaccg
     2701 gaaaactcga cgcaagaaaa atcagagaga tcctcataaa ggccaagaag ggcggaaaga
     2761 tcgccgtgta attctagcta gagaattaat tcggtacgct gaaatcacca gtctctctct
     2821 acaaatctat ctctctctat tttctccata aataatgtgt gagtagtttc ccgataaggg
     2881 aaattagggt tcttataggg tttcgctcat gtgttgagca tataagaaac ccttagtatg
     2941 tatttgtatt tgtaaaatac ttctatcaat aaaatttcta attcctaaaa ccaaaatcca
     3001 gtactaaaat ccagatcgat aacgttacac cacaatatat cctgccaaga tctcatgtga
     3061 gcaaaaggcc agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat
     3121 aggctccgcc cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac
     3181 ccgacaggac tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct
     3241 gttccgaccc tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg
     3301 ctttctcata gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg
     3361 ggctgtgtgc acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt
     3421 cttgagtcca acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg
     3481 attagcagag cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac
     3541 ggctacacta gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga
     3601 agaagagttg gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt
     3661 gtttgcaagc agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt
     3721 tctacggggt ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga
     3781 ttatcaaaaa ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc
     3841 taaagtatat atgtgtaaca ttggtctagt gattagaaaa actcatcgag catcaaatga
     3901 aactgcaatt tattcatatc aggattatca ataccatatt tttgaaaaag ccgtttctgt
     3961 aatgaaggag aaaactcacc gaggcagttc cataggatgg caagatcctg gtatcggtct
     4021 gcgattccga ctcgtccaac atcaatacaa cctattaatt tcccctcgtc aaaaataagg
     4081 ttatcaagtg agaaatcacc atgagtgacg actgaatccg gtgagaatgg caaaagttta
     4141 tgcatttctt tccagacttg ttcaacaggc cagccattac gctcgtcatc aaaatcactc
     4201 gcatcaacca aaccgttatt cattcgtgat tgcgcctgag cgagacgaaa tacgcgatcg
     4261 ctgttaaaag gacaattaca aacaggaatc gaatgcaacc ggcgcaggaa cactgccagc
     4321 gcatcaacaa tattttcacc tgaatcagga tattcttcta atacctggaa tgctgttttc
     4381 cctgggatcg cagtggtgag taaccatgca tcatcaggag tacggataaa atgcttgatg
     4441 gtcggaagag gcataaattc cgtcagccag tttagtctga ccatctcatc tgtaacatca
     4501 ttggcaacgc tacctttgcc atgtttcaga aacaactctg gcgcatcggg cttcccatac
     4561 aatcggtaga ttgtcgcacc tgattgcccg acattatcgc gagcccattt atacccatat
     4621 aaatcagcat ccatgttgga atttaatcgc ggccttgagc aagacgtttc ccgttgaata
     4681 tggctcataa caccccttgt attactgttt atgtaagcag acagttttat tgttcatgat
     4741 gatatatttt tatcttgtgc aatgtaacat cagagatttt gagacacaac gtggctttgt
     4801 tgaataaatc gaacttttgc tgagttgaag gatcagatca cgcatcttcc cgacaacgca
     4861 gaccgttccg tggcaaagca aaagttcaaa atcaccaact ggtccaccta caacaaagct
     4921 ctcatcaacc gtggctccct cactttctgg ctggatgatg gggcgattca ggcgatcccc
     4981 atccaacagc ccgccgtcga gcgggctttt ttatccccgg aagcctgtgg atagagggta
     5041 gttatccacg tgaaaccgct aatgccccgc aaagccttga ttcacggggc tttccggccc
     5101 gctccaaaaa ctatccacgt gaaatcgcta atcagggtac gtgaaatcgc taatcggagt
     5161 acgtgaaatc gctaataagg tcacgtgaaa tcgctaatca aaaaggcacg tgagaacgct
     5221 aatagccctt tcagatcaac agcttgcaaa cacccctcgc tccggcaagt agttacagca
     5281 agtagtatgt tcaattagct tttcaattat gaatatatat atcaattatt ggtcgccctt
     5341 ggcttgtgga caatgcgcta cgcgcaccgg ctccgcccgt ggacaaccgc aagcggttgc
     5401 ccaccgtcga gcgcctttgc ccacaacccg gcggccggcc gcaacagatc gttttataaa
     5461 tttttttttt tgaaaaagaa aaagcccgaa aggcggcaac ctctcgggct tctggatttc
     5521 cgatccccgg aatta
//
