LOCUS       sequence-36115-e        4657 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 4657)
  AUTHORS   Osborn MJ, Defeo AP, Blazar B, Tolar J
  TITLE     Synthetic Zinc Finger Nuclease Design and Rapid Assembly.
  JOURNAL   Hum Gene Ther. 2011 Jun 13.
  PUBMED    21663559
REFERENCE   2  (bases 1 to 4657)
  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: "Hum Gene 
            Ther. 2011 Jun 13."
COMMENT     Sequence Label: pMJO-7
FEATURES             Location/Qualifiers
     source          1..4657
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             942..1007
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1014..1034
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1691..1744
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             1751..1816
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             1823..1843
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1892..2476
                     /codon_start=1
                     /product="nonspecific DNA cleavage domain of the FokI 
                     endonuclease (Li et al., 1992)"
                     /label=FokI cleavage domain
                     /note="must dimerize to be catalytically active"
                     /translation="LVKSELEEKKSELRHKLKYVPHEYIELIEIARNSTQDRILEMKVM
                     EFFMKVYGYRGKHLGGSRKPDGAIYTVGSPIDYGVIVDTKAYSGGYNLPIGQADEMERY
                     VEENQTRNKHLNPNEWWKVYPSSVTEFKFLFVSGHFKGNYKAQLTRLNHITNCNGAVLS
                     VEELLIGGEMIKAGTLTLEEVRRKFNNGEINF"
     CDS             2498..2539
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2549..2566
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2589..2606)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2595..2819
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      complement(2902..3490)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2982..3001)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3661..4452,4453..4521))
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEADKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     primer_bind     4284..4303
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4522..4626)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtcgactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggac tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagt
      901 taagcttggt accgagctcg gatccagtac ccttcaccat ggattataag gatcacgatg
      961 gcgactacaa ggaccacgat attgactaca aagacgatga cgataaaatg gctcctaaga
     1021 aaaagcgcaa agtcggtatc catggcgttc cctctagata acgcacggac cgggtgcggc
     1081 agcccgcgcc ctggtgaagt ccgagctgga agaaaaaaag agcgagctgc gccacaagct
     1141 caagtacgtg ccccacgagt acatcgagct gatcgagatc gcccgcaaca gcacccaaga
     1201 ccgcatcctg gagatgaaag tgatggagtt cttcatgaag gtgtacggct accgcggcaa
     1261 gcacctgggc ggctcccgca agcccgatgg cgccatctac accgtgggct cccccatcga
     1321 ctatggcgtc attgtcgaca ccaaggccta ctccggcggc tacaacttac ccatcggtca
     1381 ggccgacgag atgcaacgct acgtgaagga gaaccagacc cgcaataagc acattaatcc
     1441 caacgagtgg tggaaggtgt acccctcctc cgtgaccgag ttcaaattcc tgttcgtgtc
     1501 cggccacttc aagggcaatt ataaggccca actgacccgc ctgaaccaca agaccaactg
     1561 caacggcgcc gtgctgtccg tggaggaact gctgatcggc ggcgagatga tcaaggctgg
     1621 taccctgacc ctggaagagg tgcgccgcaa gttcaacaat ggtgaaatca atttcaggtc
     1681 cggcggcgga gagggcagag gaagtcttct aacatgcggt gacgtggagg agaatcccgg
     1741 ccctaggatg gactacaaag accatgacgg tgattataaa gatcatgaca tcgattacaa
     1801 ggatgacgat gacaagatgg cccccaagaa gaagaggaag gtgggcattc acggggtgcc
     1861 ggctagctag taagcttctg cagccagagc tctagtcaaa agtgaactgg aggagaagaa
     1921 atctgaactt cgtcataaat tgaaatatgt gcctcatgaa tatattgaat taattgaaat
     1981 tgccagaaat tccactcagg atagaattct tgaaatgaag gtaatggaat tttttatgaa
     2041 agtttatgga tatagaggta aacatttggg tggatcaagg aaaccggacg gagcaattta
     2101 tactgtcgga tctcctattg attacggtgt gatcgtggat actaaagctt atagcggagg
     2161 ttataatctg ccaattggcc aagcagatga aatggagcga tatgtcgaag aaaatcaaac
     2221 acgaaacaaa catctcaacc ctaatgaatg gtggaaagtc tatccatctt ctgtaacgga
     2281 atttaagttt ttatttgtga gtggtcactt taaaggaaac tacaaagctc agcttacacg
     2341 attaaatcat atcactaatt gtaatggagc tgttcttagt gtagaagagc ttttaattgg
     2401 tggagaaatg attaaagccg gcacattaac cttagaggaa gtgagacgga aatttaataa
     2461 cggcgagata aacttttatt gagctcggcc cttcgaaggt aagcctatcc ctaaccctct
     2521 cctcggtctc gattctacgc gtaccggtca tcatcaccat caccattgag tttaaacccg
     2581 ctgatcagcc tcgactgtgc cttctagttg ccagccatct gttgtttgcc cctcccccgt
     2641 gccttccttg accctggaag gtgccactcc cactgtcctt tcctaataaa atgaggaaat
     2701 tgcatcgcat tgtctgagta ggtgtcattc tattctgggg ggtggggtgg ggcaggacag
     2761 caagggggag gattgggaag acaatagcag gcatgctggg gatgcggtgg gctctatggc
     2821 ttctgaggcg gaaagaacca gctacagagc aaaaggccag caaaaggcca ggaaccgtaa
     2881 aaaggccgcg ttgctggcgt ttttccatag gctccgcccc cctgacgagc atcacaaaaa
     2941 tcgacgctca agtcagaggt ggcgaaaccc gacaggacta taaagatacc aggcgtttcc
     3001 ccctggaagc tccctcgtgc gctctcctgt tccgaccctg ccgcttaccg gatacctgtc
     3061 cgcctttctc ccttcgggaa gcgtggcgct ttctcaatgc tcacgctgta ggtatctcag
     3121 ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga
     3181 ccgctgcgcc ttatccggta actatcgtct tgagtccaac ccggtaagac acgacttatc
     3241 gccactggca gcagccactg gtaacaggat tagcagagcg aggtatgtag gcggtgctac
     3301 agagttcttg aagtggtggc ctaactacgg ctacactaga aggacagtat ttggtatctg
     3361 cgctctgctg aagccagtta ccttcggaaa aagagttggt agctcttgat ccggcaaaca
     3421 aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa
     3481 aggatctcaa gaagatcctt tgatcttttc tacggggtct gacgctcagt ggaacgaaaa
     3541 ctcacgttaa gggattttgg tcatgagatt atcaaaaagg atcttcacct agatcctttt
     3601 aaattaaaaa tgaagtttta aatcaatcta aagtatatat gagtaaactt ggtctgacag
     3661 ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc tgtctatttc gttcatccat
     3721 agttgcctga ctccccgtcg tgtagataac tacgatacgg gagggcttac catctggccc
     3781 cagtgctgca atgataccgc gagacccacg ctcaccggct ccagatttat cagcaataaa
     3841 ccagccagcc ggaagggccg agcgcagaag tggtcctgca actttatccg cctccatcca
     3901 gtctattaat tgttgccggg aagctagagt aagtagttcg ccagttaata gtttgcgcaa
     3961 cgttgttgcc attgctacag gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt
     4021 cagctccggt tcccaacgat caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc
     4081 ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag ttggccgcag tgttatcact
     4141 catggttatg gcagcactgc ataattctct tactgtcatg ccatccgtaa gatgcttttc
     4201 tgtgactggt gagtactcaa ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg
     4261 ctcttgcccg gcgtcaatac gggataatac cgcgccacat agcagaactt taaaagtgct
     4321 catcattgga aaacgttctt cggggcgaaa actctcaagg atcttaccgc tgttgagatc
     4381 cagttcgatg taacccactc gtgcacccaa ctgatcttca gcatctttta ctttcaccag
     4441 cgtttctggg tgagcaaaaa caggaaggca aaatgccgca aaaaagggaa taagggcgac
     4501 acggaaatgt tgaatactca tactcttcct ttttcaatat tattgaagca tttatcaggg
     4561 ttattgtctc atgagcggat acatatttga atgtatttag aaaaataaac aaataggggt
     4621 tccgcgcaca tttccccgaa aagtgccacc tgacgtc
//
