LOCUS       sequence-317643-        4953 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of 3xFLAG-NLS-VP64-intN under ZF2x6-C the 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4953)
  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 4953)
  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: pJM571 ZF2x6-C 3xFLAG-NLS-VP64-intN in TUPV2
FEATURES             Location/Qualifiers
     source          1..4953
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      561..989
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(648..667)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     858..879
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1012..1208
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      1059..1194
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1121..1140
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    1328..1449
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1365..1384)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1419..1438
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1498..1514)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1498..1514)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1511..1533)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1522..1538
                     /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(1546..1552,1553..1570,1571..1576))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1591..1612
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1729..1746)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(1900..2488)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1980..1999)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2659..3450,3451..3519))
                     /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     3282..3301
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3520..3624)
                     /gene="bla"
                     /label=AmpR promoter
     regulatory      3868..3877
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3877..3942
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             3949..3969
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3985..4134
                     /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"
     primer_bind     complement(4515..4532)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4521..4745
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 acggggaagg tggcacggga tcgctttcct ctgaacgctt ctcgctgctc tttgagcctg
       61 cagacacctg gggggatacg gggaaaaagc tttaggctga aagagagatt tagaatgaca
      121 gaatcataga acggcctgcg tacggatggg gctctagctc tagagcttga tatcgaattc
      181 ctgcagcccc gctagaggga cagccccccc ccaaagcccc cagggatgta attacgtccc
      241 tcccccgcta gggggcagca gcgagccgcc cggggctccg ctccggtccg gcgctccccc
      301 cgcatccccg agccggcagc gtgcggggac agcccgggca cggggaaggt ggcacgggat
      361 cgctttcctc tgaacgcttc tcgctgctct ttgagcctgc agacacctgg ggggatacgg
      421 ggaaaaagct ttaggctgaa agagagattt agaatgacag aatcatagaa cggcctgggc
      481 gcgcctggtg gcgaacgatc aacagacggg agaggactaa agtcttcgct gacagcgacc
      541 ggactctagg gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt
      601 ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt
      661 cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct
      721 ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg
      781 tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga
      841 gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc
      901 ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga
      961 gctgatttaa caaaaattta acgcgaatta attctgtgga atgtgtgtca gttaggctca
     1021 attagtcagc aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca
     1081 gttccgccca ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg
     1141 ccgcctctgc ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct
     1201 tttgcaaaaa gctcccggga gcttgtatat ccattttcgg aggttgggct tcggaatcgt
     1261 tttccgggac gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc
     1321 ccaccccaac ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa
     1381 tttcacaaat aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa
     1441 tgtatcttat catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc
     1501 atagctgttt cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg
     1561 aagcataaag tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt
     1621 gcgctcactg cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg
     1681 ccaacgcgcg gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga
     1741 ctcgctgcgc tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat
     1801 acggttatcc acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca
     1861 aaaggccagg aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc
     1921 tgacgagcat cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata
     1981 aagataccag gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc
     2041 gcttaccgga tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc
     2101 acgctgtagg tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga
     2161 accccccgtt cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc
     2221 ggtaagacac gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag
     2281 gtatgtaggc ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag
     2341 aacagtattt ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag
     2401 ctcttgatcc ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca
     2461 gattacgcgc agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga
     2521 cgctcagtgg aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat
     2581 cttcacctag atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga
     2641 gtaaacttgg tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg
     2701 tctatttcgt tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga
     2761 gggcttacca tctggcccca gtgctgcaat gataccgcga gatccacgct caccggctcc
     2821 agatttatca gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac
     2881 tttatccgcc tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc
     2941 agttaatagt ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc
     3001 gtttggtatg gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc
     3061 catgttgtgc aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt
     3121 ggccgcagtg ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc
     3181 atccgtaaga tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg
     3241 tatgcggcga ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag
     3301 cagaacttta aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat
     3361 cttaccgctg ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc
     3421 atcttttact ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa
     3481 aaagggaata agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta
     3541 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     3601 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg
     3661 atcgggagat cgaagacggg caaaaccgat gttgacggac taatcctgac aagatctgcg
     3721 aggaggacgt gacgcgagag gacgtgtgag ctgaggacgt ggactaagag gacgtgacgc
     3781 gagaggacgt gtgagctgag gacgtgaacg ctctctagag ggtatataat gggggccact
     3841 agtctactac cagagctcat cgctagcgcc accatggact ataaagacca tgacggtgac
     3901 tacaaggatc acgacataga ttataaagac gatgacgaca agatggcacc taagaagaag
     3961 cgcaaagttg gctctggcga attcgacgcc cttgatgact ttgacttgga catgctgggg
     4021 tcagacgcac tcgatgattt tgaccttgat atgctgggaa gcgatgcact tgatgacttt
     4081 gacttggaca tgctcggctc cgacgctctt gatgatttcg atcttgacat gctcggatcc
     4141 ggtggtggcg ggtctggagg aggaggtact ggtggaggtg gcactggtac cagaagcgga
     4201 tactgcctgg acctgaagac ccaggtgcag acccctcagg gcatgaagga gatcagcaac
     4261 atccaggtgg gcgacctggt gctgagcaac accggctaca acgaggtgct gaacgtgttc
     4321 cccaagagca agaagaagag ctacaagatc accctggagg acggcaagga gatcatctgc
     4381 agcgaggagc acctgttccc cacccagacc ggcgagatga acatcagcgg cggcctgaag
     4441 gagggcatgt gcctgtacgt gaaggagtga gcggccgctc gagtctagag ggcccgttta
     4501 aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     4561 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     4621 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
     4681 ggacagcaag ggggaggatt gggaagtcaa tagcaggcat gctggggatg cggtgggctc
     4741 tatggcttct gaggcaggtc gaagaactat ttcctggctg ttacgcgagc gcatcgcgat
     4801 aagctctaga gggacagccc ccccccaaag cccccaggga tgtaattacg tccctccccc
     4861 gctagggggc agcagcgagc cgcccggggc tccgctccgg tccggcgctc cccccgcatc
     4921 cccgagccgg cagcgtgcgg ggacagcccg ggc
//
