LOCUS       sequence-335235-        3294 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 3294)
  TITLE     Plasmids to assay for nuclease cleavage in planta
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 3294)
  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            "Unpublished"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 0000 (No LUC)
FEATURES             Location/Qualifiers
     source          1..3294
                     /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"
     CDS             403..768
                     /codon_start=1
                     /gene="lacZ fragment"
                     /product="LacZ-alpha fragment of beta-galactosidase"
                     /label=lacZ-alpha
                     /translation="MTMITPSAQLTLTKGNKSWSSTAVAAALELVDPPGCRNSISSLSI
                     PSTSRGGPVPNSPYSESYYARSLAVVLQRRDWENPGVTQLNRLAAHPPFASWRNSEEAR
                     TDRPSQQLRSLNGEWKL"
     primer_bind     426..446
                     /label=T3
                     /note="T3 promoter, forward primer"
     promoter        428..446
                     /label=T3 promoter
                     /note="promoter for bacteriophage T3 RNA polymerase"
     misc_feature    459..566
                     /label=MCS
                     /note="pBluescript multiple cloning site"
     primer_bind     483..499
                     /label=SK primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     483..499
                     /label=pBluescriptSK
                     /note="For pBluescript vector"
     primer_bind     complement(533..549)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(534..550)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     complement(574..593)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(575..593)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(603..620)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(603..619)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(612..634)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     misc_feature    784..806
                     /label=LB T-DNA repeat
                     /note="left border repeat from nopaline C58 T-DNA 
                     (truncated)"
     rep_origin      933..1368
                     /label=pSa ori
                     /note="origin of replication from bacterial plasmid pSa"
     primer_bind     complement(1648..1667)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1659..2474
                     /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             1659..2474
                     /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     complement(1732..1751)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     rep_origin      2645..3233
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     3134..3153
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
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 gctctagaac tagtggatcc cccgggctgc aggaattcga tatcaagctt atcgataccg
      541 tcgacctcga gggggggccc ggtacccaat tcgccctata gtgagtcgta ttacgcgcgc
      601 tcactggccg tcgttttaca acgtcgtgac tgggaaaacc ctggcgttac ccaacttaat
      661 cgccttgcag cacatccccc tttcgccagc tggcgtaata gcgaagaggc ccgcaccgat
      721 cgcccttccc aacagttgcg cagcctgaat ggcgaatgga aattgtaagc gttaatgtta
      781 acgttacacc acaatatatc ctgccaagat ctaattccgg ggatcggaaa tccagaagcc
      841 cgagaggttg ccgcctttcg ggctttttct ttttcaaaaa aaaaaattta taaaacgatc
      901 tgttgcggcc ggccgccggg ttgtgggcaa aggcgctcga cggtgggcaa ccgcttgcgg
      961 ttgtccacgg gcggagccgg tgcgcgtagc gcattgtcca caagccaagg gcgaccaata
     1021 attgatatat atattcataa ttgaaaagct aattgaacat actacttgct gtaactactt
     1081 gccggagcga ggggtgtttg caagctgttg atctgaaagg gctattagcg ttctcacgtg
     1141 cctttttgat tagcgatttc acgtgacctt attagcgatt tcacgtactc cgattagcga
     1201 tttcacgtac cctgattagc gatttcacgt ggatagtttt tggagcgggc cggaaagccc
     1261 cgtgaatcaa ggctttgcgg ggcattagcg gtttcacgtg gataactacc ctctatccac
     1321 aggcttccgg ggataaaaaa gcccgctcga cggcgggctg ttggatgggg atcgcctgaa
     1381 tcgccccatc atccagccag aaagtgaggg agccacggtt gatgagagct ttgttgtagg
     1441 tggaccagtt ggtgattttg aacttttgct ttgccacgga acggtctgcg ttgtcgggaa
     1501 gatgcgtgat ctgatccttc aactcagcaa aagttcgatt tattcaacaa agccacgttg
     1561 tgtctcaaaa tctctgatgt tacattgcac aagataaaaa tatatcatca tgaacaataa
     1621 aactgtctgc ttacataaac agtaatacaa ggggtgttat gagccatatt caacgggaaa
     1681 cgtcttgctc aaggccgcga ttaaattcca acatggatgc tgatttatat gggtataaat
     1741 gggctcgcga taatgtcggg caatcaggtg cgacaatcta ccgattgtat gggaagcccg
     1801 atgcgccaga gttgtttctg aaacatggca aaggtagcgt tgccaatgat gttacagatg
     1861 agatggtcag actaaactgg ctgacggaat ttatgcctct tccgaccatc aagcatttta
     1921 tccgtactcc tgatgatgca tggttactca ccactgcgat cccagggaaa acagcattcc
     1981 aggtattaga agaatatcct gattcaggtg aaaatattgt tgatgcgctg gcagtgttcc
     2041 tgcgccggtt gcattcgatt cctgtttgta attgtccttt taacagcgat cgcgtatttc
     2101 gtctcgctca ggcgcaatca cgaatgaata acggtttggt tgatgcgagt gattttgatg
     2161 acgagcgtaa tggctggcct gttgaacaag tctggaaaga aatgcataaa cttttgccat
     2221 tctcaccgga ttcagtcgtc actcatggtg atttctcact tgataacctt atttttgacg
     2281 aggggaaatt aataggttgt attgatgttg gacgagtcgg aatcgcagac cgataccagg
     2341 atcttgccat cctatggaac tgcctcggtg agttttctcc ttcattacag aaacggcttt
     2401 ttcaaaaata tggtattgat aatcctgata tgaataaatt gcagtttcat ttgatgctcg
     2461 atgagttttt ctaatcacta gaccaatgtt acacatatat actttagatt gatttaaaac
     2521 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     2581 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     2641 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     2701 taccagcggt ggtttgtttg ccggatcaag agctaccaac tcttcttccg aaggtaactg
     2761 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     2821 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     2881 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     2941 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     3001 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     3061 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     3121 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     3181 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     3241 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atga
//
