LOCUS       sequence-318788-        5187 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of 3xFLAG-NLS-VP64-intC-ZF1-NLS-HA under the 
            ZF2x6-C promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5187)
  AUTHORS   Muldoon JJ, Kandula V, Hong M, Donahue PS, Boucher JD, Bagheri N, 
            Leonard JN
  TITLE     Model-guided design of mammalian genetic programs.
  JOURNAL   Sci Adv. 2021 Feb 19;7(8). pii: 7/8/eabe9375. doi: 
            10.1126/sciadv.abe9375. Print 2021 Feb.
  PUBMED    33608279
REFERENCE   2  (bases 1 to 5187)
  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: "Sci Adv."; 
            date: "2021-02-19"; volume: "7(8). pii: 7/8/eabe9375. doi"; pages: "
            10.1126/sciadv.abe9375. Print 2021 Feb"
COMMENT     Sequence Label: pJM576 ZF2x6-C 3xFLAG-NLS-VP64-intC-ZF1-NLS-HA...
FEATURES             Location/Qualifiers
     source          1..5187
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        complement(1..105)
                     /gene="bla"
                     /label=AmpR promoter
     regulatory      349..358
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             358..423
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             430..450
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             466..615
                     /codon_start=1
                     /product="tetrameric repeat of the minimal activation 
                     domain of herpes simplex virus VP16 (Beerli et al., 1998)"
                     /label=VP64
                     /translation="DALDDFDLDMLGSDALDDFDLDMLGSDALDDFDLDMLGSDALDDF
                     DLDML"
     CDS             1123..1143
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1156..1182
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(1230..1247)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1236..1460
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2229..2657
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2316..2335)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2526..2547
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2680..2876
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      2727..2862
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2789..2808
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    2996..3117
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3033..3052)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3087..3106
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3166..3182)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3166..3182)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3179..3201)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3190..3206
                     /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(3214..3220,3221..3238,3239..3244))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3259..3280
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3397..3414)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3568..4156)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3648..3667)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4327..5118,5119..5187))
                     /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     4950..4969
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 actcttcctt tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata
       61 catatttgaa tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa
      121 agtgccacct gacgtcgacg gatcgggaga tcgaagacgg gcaaaaccga tgttgacgga
      181 ctaatcctga caagatctgc gaggaggacg tgacgcgaga ggacgtgtga gctgaggacg
      241 tggactaaga ggacgtgacg cgagaggacg tgtgagctga ggacgtgaac gctctctaga
      301 gggtatataa tgggggccac tagtctacta ccagagctca tcgctagcgc caccatggac
      361 tataaagacc atgacggtga ctacaaggat cacgacatag attataaaga cgatgacgac
      421 aagatggcac ctaagaagaa gcgcaaagtt ggctctggcg aattcgacgc ccttgatgac
      481 tttgacttgg acatgctggg gtcagacgca ctcgatgatt ttgaccttga tatgctggga
      541 agcgatgcac ttgatgactt tgacttggac atgctcggct ccgacgctct tgatgatttc
      601 gatcttgaca tgctcggatc cggtggtggc gggtctggtg gaggaggtag tggtggaggt
      661 ggctctgcca ccatgatgct gaagaagatc ctgaagatcg aggagctgga cgagagagag
      721 ctgatcgaca tcgaggtgag cggcaaccac ctgttctacg ccaacgacat cctgacccac
      781 aacagcaaca ggggtggtgg cgggtctggt ggaggaggta gtggtggagg tggctctggt
      841 accgctagac ccggggagcg ccccttccag tgtcgcattt gcatgcggaa cttttcgcgc
      901 caggacaggc ttgacaggca tacccgtact cataccggtg aaaaaccgtt tcagtgtcgg
      961 atctgtatgc gaaatttctc ccagaaggag cacttggcgg ggcatctacg tacgcacacc
     1021 ggcgagaagc cattccaatg ccgaatatgc atgcgcaact tcagtcgccg cgacaacctg
     1081 aaccggcacc taaaaaccca cctgagagaa ttcggctctg gccctaagaa gaagcgcaaa
     1141 gttggctctg gcatgtaccc atacgacgta ccagattacg cttaggcggc cgctcgagtc
     1201 tagagggccc gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
     1261 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
     1321 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
     1381 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gtcaatagca ggcatgctgg
     1441 ggatgcggtg ggctctatgg cttctgaggc aggtcgaaga actatttcct ggctgttacg
     1501 cgagcgcatc gcgataagct ctagagggac agcccccccc caaagccccc agggatgtaa
     1561 ttacgtccct cccccgctag ggggcagcag cgagccgccc ggggctccgc tccggtccgg
     1621 cgctcccccc gcatccccga gccggcagcg tgcggggaca gcccgggcac ggggaaggtg
     1681 gcacgggatc gctttcctct gaacgcttct cgctgctctt tgagcctgca gacacctggg
     1741 gggatacggg gaaaaagctt taggctgaaa gagagattta gaatgacaga atcatagaac
     1801 ggcctgcgta cggatggggc tctagctcta gagcttgata tcgaattcct gcagccccgc
     1861 tagagggaca gccccccccc aaagccccca gggatgtaat tacgtccctc ccccgctagg
     1921 gggcagcagc gagccgcccg gggctccgct ccggtccggc gctccccccg catccccgag
     1981 ccggcagcgt gcggggacag cccgggcacg gggaaggtgg cacgggatcg ctttcctctg
     2041 aacgcttctc gctgctcttt gagcctgcag acacctgggg ggatacgggg aaaaagcttt
     2101 aggctgaaag agagatttag aatgacagaa tcatagaacg gcctgggcgc gcctggtggc
     2161 gaacgatcaa cagacgggag aggactaaag tcttcgctga cagcgaccgg actctagggg
     2221 gtatccccac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg ttacgcgcag
     2281 cgtgaccgct acacttgcca gcgccctagc gcccgctcct ttcgctttct tcccttcctt
     2341 tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc ctttagggtt
     2401 ccgatttagt gctttacggc acctcgaccc caaaaaactt gattagggtg atggttcacg
     2461 tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt ccacgttctt
     2521 taatagtgga ctcttgttcc aaactggaac aacactcaac cctatctcgg tctattcttt
     2581 tgatttataa gggattttgc cgatttcggc ctattggtta aaaaatgagc tgatttaaca
     2641 aaaatttaac gcgaattaat tctgtggaat gtgtgtcagt taggctcaat tagtcagcaa
     2701 ccatagtccc gcccctaact ccgcccatcc cgcccctaac tccgcccagt tccgcccatt
     2761 ctccgcccca tggctgacta atttttttta tttatgcaga ggccgaggcc gcctctgcct
     2821 ctgagctatt ccagaagtag tgaggaggct tttttggagg cctaggcttt tgcaaaaagc
     2881 tcccgggagc ttgtatatcc attttcggag gttgggcttc ggaatcgttt tccgggacgc
     2941 cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc accccaactt
     3001 gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa
     3061 agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca
     3121 tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat agctgtttcc
     3181 tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa gcataaagtg
     3241 taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc gctcactgcc
     3301 cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc aacgcgcggg
     3361 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     3421 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     3481 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     3541 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     3601 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     3661 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     3721 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     3781 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     3841 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     3901 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     3961 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     4021 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     4081 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     4141 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     4201 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     4261 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     4321 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     4381 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     4441 tggccccagt gctgcaatga taccgcgaga tccacgctca ccggctccag atttatcagc
     4501 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     4561 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     4621 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     4681 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     4741 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     4801 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     4861 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     4921 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     4981 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     5041 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     5101 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     5161 ggcgacacgg aaatgttgaa tactcat
//
