LOCUS       Exported                5475 bp ds-DNA     circular SYN 16-SEP-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5475)
  AUTHORS   NIH-NCI
  TITLE     Direct Submission
  JOURNAL   Exported Tuesday, Sep 16, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5475
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     polyA_signal    390..511
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(560..576)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     protein_bind    584..600
                     /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(608..638)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    653..674
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     rep_origin      complement(962..1550)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     CDS             complement(1721..2581)
                     /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"
     promoter        complement(2582..2686)
                     /gene="bla"
                     /label=AmpR promoter
     enhancer        2952..3331
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        3332..3535
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     protein_bind    3537..3555
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3558..3576
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3753..3777
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    3780..3971
                     /label=ZNF207 ZnF (1-64)
     CDS             3783..3971
                     /codon_start=1
                     /label=ZNF207 cDNA
                     /translation="GRKKKKQLKPWCWYCNRDFDDEKILIQHQKAKHFKCHICHKKLYT
                     GPGLAIHCMQVHKETIDA"
     misc_feature    3810..3953
                     /label=ZnF domain
     protein_bind    complement(3973..3997)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             4038..4103
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     polyA_signal    4150..4374
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    4658..4705
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             join(4713..5475,1..260)
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             join(4713..5475,1..260)
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
ORIGIN
        1 ttgcaggatc gccgcggctc cgggcgtata tgctccgcat tggtcttgac caactctatc
       61 agagcttggt tgacggcaat ttcgatgatg cagcttgggc gcagggtcga tgcgacgcaa
      121 tcgtccgatc cggagccggg actgtcgggc gtacacaaat cgcccgcaga agcgcggccg
      181 tctggaccga tggctgtgta gaagtactcg ccgatagtgg aaaccgacgc cccagcactc
      241 gtccgagggc aaaggaatag cacgtactac gagatttcga ttccaccgcc gccttctatg
      301 aaaggttggg cttcggaatc gttttccggg acgccggctg gatgatcctc cagcgcgggg
      361 atctcatgct ggagttcttc gcccacccca acttgtttat tgcagcttat aatggttaca
      421 aataaagcaa tagcatcaca aatttcacaa ataaagcatt tttttcactg cattctagtt
      481 gtggtttgtc caaactcatc aatgtatctt atcatgtctg tataccgtcg acctctagct
      541 agagcttggc gtaatcatgg tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa
      601 ttccacacaa catacgagcc ggaagcataa agtgtaaagc ctggggtgcc taatgagtga
      661 gctaactcac attaattgcg ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt
      721 gccagctgca ttaatgaatc ggccaacgcg cggggagagg cggtttgcgt attgggcgct
      781 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
      841 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
      901 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
      961 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
     1021 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
     1081 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
     1141 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
     1201 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
     1261 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
     1321 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
     1381 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
     1441 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
     1501 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
     1561 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
     1621 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
     1681 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
     1741 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     1801 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     1861 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     1921 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     1981 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     2041 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     2101 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     2161 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     2221 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     2281 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     2341 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     2401 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     2461 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     2521 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     2581 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     2641 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     2701 aagtgccacc tgacgtcgac ggatcgggag atctcccgat cccctatggt gcactctcag
     2761 tacaatctgc tctgatgccg catagttaag ccagtatctg ctccctgctt gtgtgttgga
     2821 ggtcgctgag tagtgcgcga gcaaaattta agctacaaca aggcaaggct tgaccgacaa
     2881 ttgcatgaag aatctgctta gggttaggcg ttttgcgctg cttcgcgatg tacgggccag
     2941 atatacgcgt tgacattgat tattgactag ttattaatag taatcaatta cggggtcatt
     3001 agttcatagc ccatatatgg agttccgcgt tacataactt acggtaaatg gcccgcctgg
     3061 ctgaccgccc aacgaccccc gcccattgac gtcaataatg acgtatgttc ccatagtaac
     3121 gccaataggg actttccatt gacgtcaatg ggtggagtat ttacggtaaa ctgcccactt
     3181 ggcagtacat caagtgtatc atatgccaag tacgccccct attgacgtca atgacggtaa
     3241 atggcccgcc tggcattatg cccagtacat gaccttatgg gactttccta cttggcagta
     3301 catctacgta ttagtcatcg ctattaccat ggtgatgcgg ttttggcagt acatcaatgg
     3361 gcgtggatag cggtttgact cacggggatt tccaagtctc caccccattg acgtcaatgg
     3421 gagtttgttt tggcaccaaa atcaacggga ctttccaaaa tgtcgtaaca actccgcccc
     3481 attgacgcaa atgggcggta ggcgtgtacg gtgggaggtc tatataagca gagctctccc
     3541 tatcagtgat agagatctcc ctatcagtga tagagatcgt cgacgagctc gtttagtgaa
     3601 ccgtcagatc gcctggagac gccatccacg ctgttttgac ctccatagaa gacaccggga
     3661 ccgatccagc ctccggactc tagcgtttaa acttaagctt ggtaccgagc tcggatccac
     3721 tagtccagtg tggtggaatt ctgcagatat caacaagttt gtacaaaaaa gcaggcttca
     3781 tgggtcgcaa gaagaagaag cagctgaagc cgtggtgctg gtattgtaat agagattttg
     3841 atgatgagaa gatccttatt cagcaccaaa aagcaaagca ttttaaatgc catatatgtc
     3901 acaagaaatt gtatacagga cctggcttag ctattcattg catgcaggta cataaagaaa
     3961 caatagatgc caacccagct ttcttgtaca aagtggtgat gggctgcagg aattcgatat
     4021 caagcttagg tggaatggac tacaaggacc acgacggtga ctacaaggac cacgacatcg
     4081 actacaagga cgacgacgac aagtagctcg agtctagagg gcccgtttaa acccgctgat
     4141 cagcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt
     4201 ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat
     4261 cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg
     4321 gggaggattg ggaagacaat agcaggcatg ctggggatgc ggtgggctct atggcttctg
     4381 aggcggaaag aaccagctgg ggctctaggg ggtatcccca cgcgccctgt agcggcgcat
     4441 taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccctag
     4501 cgcccgctcc tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc
     4561 aagctctaaa tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc
     4621 ccaaaaaact tgattagggt gatggttcac gtacctagaa gttcctattc cgaagttcct
     4681 attctctaga aagtatagga acttccttgg ccaaaaagcc tgaactcacc gcgacgtctg
     4741 tcgagaagtt tctgatcgaa aagttcgaca gcgtctccga cctgatgcag ctctcggagg
     4801 gcgaagaatc tcgtgctttc agcttcgatg taggagggcg tggatatgtc ctgcgggtaa
     4861 atagctgcgc cgatggtttc tacaaagatc gttatgttta tcggcacttt gcatcggccg
     4921 cgctcccgat tccggaagtg cttgacattg gggaattcag cgagagcctg acctattgca
     4981 tctcccgccg tgcacagggt gtcacgttgc aagacctgcc tgaaaccgaa ctgcccgctg
     5041 ttctgcagcc ggtcgcggag gccatggatg cgatcgctgc ggccgatctt agccagacga
     5101 gcgggttcgg cccattcgga ccgcaaggaa tcggtcaata cactacatgg cgtgatttca
     5161 tatgcgcgat tgctgatccc catgtgtatc actggcaaac tgtgatggac gacaccgtca
     5221 gtgcgtccgt cgcgcaggct ctcgatgagc tgatgctttg ggccgaggac tgccccgaag
     5281 tccggcacct cgtgcacgcg gatttcggct ccaacaatgt cctgacggac aatggccgca
     5341 taacagcggt cattgactgg agcgaggcga tgttcgggga ttcccaatac gaggtcgcca
     5401 acatcttctt ctggaggccg tggttggctt gtatggagca gcagacgcgc tacttcgagc
     5461 ggaggcatcc ggagc
//
