LOCUS       sequence-393005-        5658 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 5658)
  AUTHORS   Maeder ML, Thibodeau-Beganny S, Osiak A, Wright DA, Anthony RM, 
            Eichtinger M, Jiang T, Foley JE, Winfrey RJ, Townsend JA, 
            Unger-Wallace E, Sander JD, Muller-Lerch F, Fu F, Pearlberg J, 
            Goebel C, Dassie JP, Pruett-Miller SM, Porteus MH, Sgroi DC, Iafrate
            AJ, Dobbs D, McCray PB, Cathomen T, Voytas DF, Joung JK
  TITLE     Rapid ""open-source"" engineering of customized zinc-finger 
            nucleases for highly efficient gene modification.
  JOURNAL   Mol Cell. 2008 Jul 25. 31(2):294-301.
  PUBMED    18657511
REFERENCE   2  (bases 1 to 5658)
  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: "Mol Cell. 
            2008 Jul 25. 31(2):294-301."
COMMENT     Sequence Label: dopamine beta hydroxylase (dbh)_R (OZ526)
FEATURES             Location/Qualifiers
     source          1..5658
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        join(12..17,18..35,36..42)
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     protein_bind    50..66
                     /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)."
     CDS             368..391
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     rep_origin      793..1221
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(880..899)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1090..1111
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1555..1574)
                     /label=pENTR-R
                     /note="pENTR vectors, reverse primer"
     CDS             1566..2381
                     /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             1566..2381
                     /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(1639..1658)
                     /label=Kan-R
                     /note="Kanamycin resistance gene, reverse primer"
     primer_bind     complement(3905..3921)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    3929..3945
                     /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(3953..3959,3960..3977,3978..3983))
                     /label=lac UV5 promoter
                     /note="E. coli lac promoter with an ""up"" mutation"
     primer_bind     complement(4227..4245)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     rep_origin      4336..4924
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4825..4844
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5078..5095
                     /label=L4440
                     /note="L4440 vector, forward primer"
     misc_feature    5110..5250
                     /label=bom
                     /note="basis of mobility region from pBR322"
     primer_bind     complement(5176..5195)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5336..5358
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     CDS             complement(5352..5543)
                     /codon_start=1
                     /gene="rop"
                     /product="Rop protein, which maintains plasmids at low copy
                     number"
                     /label=rop
                     /translation="MTKQEKTALNMARFIRSQTLTLLEKLNELDADEQADICESLHDHA
                     DELYRSCLARFGDDGENL"
ORIGIN
        1 gaattccggg ctttacactt tatgcttccg gctcgtataa tgtgtcgact tgtgagcgga
       61 taacaatttc acacaggaaa cagctatgcc tcaacagcag caaatgcaac ctcccaattc
      121 aagtgcggac aacaaccctt tgcaacagca atcatcacaa aataccgtac caaacgtcct
      181 caaccaaatt aaccaaatct tttctccaga ggagcaacgc agcttattac aagaagccat
      241 cgaaacctgc aagaattttg aaaaaacaca attgggtagt acgatgacgg aacctgtcaa
      301 gcaaagtttt attaggaaat acattgtgca aaaggccctg agaaaaatcc aagctttggc
      361 ggccgctgac tacaaagacg atgatgacaa atctagaccc ggggagcgcc ccttccagtg
      421 tcgcatttgc atgcggaact tttcgacgaa cagcatgctt gccaggcata cccgtactca
      481 taccggtgaa aaaccgtttc agtgtcggat ctgtatgcga aatttctccc agaacgagca
      541 cttgaaggtc catctacgta cgcacaccgg cgagaagcca ttccaatgcc gaatatgcat
      601 gcgcaacttc agtcaggggg ggacgctgcg gcgccaccta aaaacccacc tgaggggatc
      661 ctctgcacag tagtaactag agactagaaa aaggccgaca agtcccgctc cgctgaagat
      721 cctggcgtaa tagcgaagag gcccgcaccg atcgcccttc ccaacagttg cgcagcctga
      781 atggcgaatg ggacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc
      841 gcagcgtgac cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt
      901 cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag
      961 ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgattag ggtgatggtt
     1021 cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt
     1081 tctttaatag tggactcttg ttccaaactg gaacaacact caaccctatc tcggtctatt
     1141 cttttgattt ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt
     1201 aacaaaaatt taacgcgaat tttaacaaaa tctagcgctg aggtctgcct cgtgaagaag
     1261 gtgttgctga ctcataccag gcctgaatcg ccccatcatc cagccagaaa gtgagggagc
     1321 cacggttgat gagagctttg ttgtaggtgg accagttggt gattttgaac ttttgctttg
     1381 ccacggaacg gtctgcgttg tcgggaagat gcgtgatctg atccttcaac tcagcaaaag
     1441 ttcgatttat tcaacaaagc cacgttgtgt ctcaaaatct ctgatgttac attgcacaag
     1501 ataaaaatat atcatcatga acaataaaac tgtctgctta cataaacagt aatacaaggg
     1561 gtgttatgag ccatattcaa cgggaaacgt cttgctcgag gccgcgatta aattccaaca
     1621 tggatgctga tttatatggg tataaatggg ctcgcgataa tgtcgggcaa tcaggtgcga
     1681 caatctatcg attgtatggg aagcccgatg cgccagagtt gtttctgaaa catggcaaag
     1741 gtagcgttgc caatgatgtt acagatgaga tggtcagact aaactggctg acggaattta
     1801 tgcctcttcc gaccatcaag cattttatcc gtactcctga tgatgcatgg ttactcacca
     1861 ctgcgatccc cgggaaaaca gcattccagg tattagaaga atatcctgat tcaggtgaaa
     1921 atattgttga tgcgctggca gtgttcctgc gccggttgca ttcgattcct gtttgtaatt
     1981 gtccttttaa cagcgatcgc gtatttcgtc tcgctcaggc gcaatcacga atgaataacg
     2041 gtttggttga tgcgagtgat tttgatgacg agcgtaatgg ctggcctgtt gaacaagtct
     2101 ggaaagaaat gcataagctt ttgccattct caccggattc agtcgtcact catggtgatt
     2161 tctcacttga taaccttatt tttgacgagg ggaaattaat aggttgtatt gatgttggac
     2221 gagtcggaat cgcagaccga taccaggatc ttgccatcct atggaactgc ctcggtgagt
     2281 tttctccttc attacagaaa cggctttttc aaaaatatgg tattgataat cctgatatga
     2341 ataaattgca gtttcatttg atgctcgatg agtttttcta atcagaattg gttaattggt
     2401 tgtaacactg gcagagcatt acgctgactt gacgggacgg cggctttgtt gaataaatcg
     2461 aacttttgct gagttgaagg atcagatcac gcatcttccc gacaacgcag accgttccgt
     2521 ggcaaagcaa aagttcaaaa tcaccaactg gtccacctac aacaaagctc tcatcaaccg
     2581 tggctccctc actttctggc tggatgatgg ggcgattcag gcctggtatg agtcagcaac
     2641 accttcttca cgaggcagac ctcagcgctc aaagatgcag gggtaaaagc taaccgcatc
     2701 tttaccgaca aggcatccgg cagttcaaca gatcgggaag ggctggattt gctgaggatg
     2761 aaggtggagg aaggtgatgt cattctggtg aagaagctcg accgtcttgg ccgcgacacc
     2821 gccgacatga tccaactgat aaaagagttt gatgctcagg gtgtagcggt tcggtttatt
     2881 gacgacggga tcagtaccga cggtgatatg gggcaaatgg tggtcaccat cctgtcggct
     2941 gtggcacagg ctgaacgccg gaggatcagc catgctaggt caaaataatc ctgttaacaa
     3001 tgcttttata tcctgtaaag aatccatttt caaaatcatg tcaaggtcct cccgaggaaa
     3061 aatcagtaga aatagctgtt ccagtctttc tagccttgat tctgcggccg cctctggttt
     3121 ctcttctttc acttcaggct gacgtacatc acgtaagtca acgaaagctt ccagttgttc
     3181 agccagaatg gttgccgcac gacgaatcgc ctcttcagga tcgattgtgc cgttggtttc
     3241 catttcgatg accagcttgt ccaggtcggt acgctgttct acacgcgctg cttcaacatt
     3301 gtaggcaata cgctccacag ggctgtagca tgcgtcgacc agcagacggc cgattgggcg
     3361 ctcatcttct tccgaatgaa ttcgggtaga agccggcaca taaccacgac cgcgctgaac
     3421 tttgatacgc atgctaatag acgcgttctc atcggtcagg tggcagatca cgtgctgcgg
     3481 cttgacgatt tcgacatcac cgtcgtgggt gatatcggct gcagtcacag ggccaatgcc
     3541 agatttattc aaggtaagaa taacttcatc tttgccctga actctcaccg ccagcccttt
     3601 caggttgagc aggatttcca ggatatcttc ctgaacgcct tctttggtgc tgtactcatg
     3661 tagtacacca tcaatctcaa cctcggtcac cgcgcaaccc ggcatcgatg agagcagaat
     3721 acggcgcagt gcgttaccca gagtatggcc aaagccacgc tctaaaggct caagggtcac
     3781 cttggcgtgc gtcgaactca cttgctcgat atcaaccagg cgcggtttta gaaactctgt
     3841 cacagaaccc tgcattgtgt cctctctttg gtctagctag agttccagtg aatccgtaat
     3901 catggtcata gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacattatac
     3961 gagccggaag cataaagtgt aaagcctctc tagctagatt gagttaatct ccatgtagcg
     4021 ttacaagatc tgaaggggtt cgaattctat ttttttgatg ggaaggcact tatttttgat
     4081 cgttcgctca aagaagcatc gaacgctggg ttttagataa aaaaattgag agaaaacagg
     4141 atgcttccat cggattcatc ttagataaag tgagattata tagtgaaatc cgatttgatt
     4201 ttttattgct tctggttatc caaagggcct cgtgatacgc ctatttttat aggttaatgt
     4261 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     4321 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     4381 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     4441 gaaggtaact ggcttcagca gagcgcagat accaaatact gtccttctag tgtagccgta
     4501 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     4561 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     4621 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     4681 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     4741 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     4801 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     4861 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     4921 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     4981 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     5041 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
     5101 ggaagagcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt cacaccgcat
     5161 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagt atacactccg
     5221 ctatcgctac gtgactgggt catggctgcg ccccgacacc cgccaacacc cgctgacgcg
     5281 ccctgacggg cttgtctgct cccggcatcc gcttacagac aagctgtgac cgtctccggg
     5341 agctgcatgt gtcagaggtt ttcaccgtca tcaccgaaac gcgcgaggca gctgcggtaa
     5401 agctcatcag cgtggtcgtg aagcgattca cagatgtctg cctgttcatc cgcgtccagc
     5461 tcgttgagtt tctccagaag cgttaatgtc tggcttctga taaagcgggc catgttaagg
     5521 gcggtttttt cctgtttggt cactgatgcc tccgtgtaag ggggatttct gttcatgggg
     5581 gtaatgatac cgatgaaacg agagaggatg ctcacgatac gggttactga tgatgaacat
     5641 gcccggttac tggaacgg
//
