LOCUS       sequence-318358-        5124 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Constitutive expression of 3xFLAG-NLS-FKBP-ZF2 under the CMV 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5124)
  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 5124)
  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: pJB001 CMV 3xFLAG-NLS-FKBP-ZF2
FEATURES             Location/Qualifiers
     source          1..5124
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(33..52)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        224..603
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        604..807
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     758..778
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     852..871
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        852..870
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      895..904
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             904..969
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             976..996
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(1039..1041,1042..1362)
                     /codon_start=1
                     /product="human FK506-binding protein FKBP12"
                     /label=FKBP
                     /translation="MGVQVETISPGDGRTFPKRGQTCVVHYTGMLEDGKKFDSSRDRNK
                     PFKFMLGKQEVIRGWEEGVAQMSVGQRAKLTISPDYAYGATGHPGIIPPHATLVFDVEL
                     LKLE"
     primer_bind     complement(1743..1760)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1749..1973
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2019..2447
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2106..2125)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2316..2337
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2470..2666
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      2517..2652
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2579..2598
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    2786..2907
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2823..2842)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2877..2896
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2956..2972)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2956..2972)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2969..2991)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2980..2996
                     /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(3004..3010,3011..3028,3029..3034))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3049..3070
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3187..3204)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3358..3946)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3438..3457)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4117..4908,4909..4977))
                     /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     4740..4759
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4978..5082)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 agatctcccg atcccctatg gtgcactctc agtacaatct gctctgatgc cgcatagtta
       61 agccagtatc tgctccctgc ttgtgtgttg gaggtcgctg agtagtgcgc gagcaaaatt
      121 taagctacaa caaggcaagg cttgaccgac aattgcatga agaatctgct tagggttagg
      181 cgttttgcgc tgcttcgcga tgtacgggcc agatatacgc gttgacattg attattgact
      241 agttattaat agtaatcaat tacggggtca ttagttcata gcccatatat ggagttccgc
      301 gttacataac ttacggtaaa tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg
      361 acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca ttgacgtcaa
      421 tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta tcatatgcca
      481 agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta tgcccagtac
      541 atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat cgctattacc
      601 atggtgatgc ggttttggca gtacatcaat gggcgtggat agcggtttga ctcacgggga
      661 tttccaagtc tccaccccat tgacgtcaat gggagtttgt tttggcacca aaatcaacgg
      721 gactttccaa aatgtcgtaa caactccgcc ccattgacgc aaatgggcgg taggcgtgta
      781 cggtgggagg tctatataag cagagctctc tggctaacta gagaacccac tgcttactgg
      841 cttatcgaaa ttaatacgac tcactatagg gagtcccaag ctggctagct taaggccacc
      901 atggattata aggatcacga tggcgactac aaggaccacg atattgacta caaagacgat
      961 gacgataaaa tggctcctaa gaaaaagcgc aaagtcggta tccatggcgt tcccgtcgac
     1021 ggtggactcc aagttaagat gggagtgcag gtggaaacca tctccccagg agacgggcgc
     1081 accttcccca agcgcggcca gacctgcgtg gtgcactaca ccgggatgct tgaagatgga
     1141 aagaaatttg attcctcccg ggacagaaac aagcccttta agtttatgct aggcaagcag
     1201 gaggtgatcc gaggctggga agaaggggtt gcccagatga gtgtgggtca gagagccaaa
     1261 ctgactatat ctccagatta tgcctatggt gccactgggc acccaggcat catcccacca
     1321 catgccactc tcgtctttga tgtggagctt ctaaaactgg aaggatccgg tggtggcggg
     1381 tctggaggag gaggtagtgg tggaggtggc tctggtaccg gtggagctag acccggggag
     1441 cgccccttcc agtgtcgcat ttgcatgcgg aacttttcga gaaatttcat tttgcaaaga
     1501 catacccgta ctcataccgg tgaaaaaccg tttcagtgtc ggatctgtat gcgaaatttc
     1561 tccgatagag ctaatttgag gaggcatctg cgtacgcaca ccggcgagaa gccattccaa
     1621 tgccgaatat gcatgcgcaa cttcagtaga catgatcaat tgactagaca cctaaaaacc
     1681 cacctgagag gatcttaagc ggccgctcga gtctagaggg cccgtttaaa cccgctgatc
     1741 agcctcgact gtgccttcta gttgccagcc atctgttgtt tgcccctccc ccgtgccttc
     1801 cttgaccctg gaaggtgcca ctcccactgt cctttcctaa taaaatgagg aaattgcatc
     1861 gcattgtctg agtaggtgtc attctattct ggggggtggg gtggggcagg acagcaaggg
     1921 ggaggattgg gaagtcaata gcaggcatgc tggggatgcg gtgggctcta tggcttctga
     1981 ggcggaaaga accagctggg gctctagggg gtatccccac gcgccctgta gcggcgcatt
     2041 aagcgcggcg ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccctagc
     2101 gcccgctcct ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca
     2161 agctctaaat cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc
     2221 caaaaaactt gattagggtg atggttcacg tagtgggcca tcgccctgat agacggtttt
     2281 tcgccctttg acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac
     2341 aacactcaac cctatctcgg tctattcttt tgatttataa gggattttgc cgatttcggc
     2401 ctattggtta aaaaatgagc tgatttaaca aaaatttaac gcgaattaat tctgtggaat
     2461 gtgtgtcagt taggctcaat tagtcagcaa ccatagtccc gcccctaact ccgcccatcc
     2521 cgcccctaac tccgcccagt tccgcccatt ctccgcccca tggctgacta atttttttta
     2581 tttatgcaga ggccgaggcc gcctctgcct ctgagctatt ccagaagtag tgaggaggct
     2641 tttttggagg cctaggcttt tgcaaaaagc tcccgggagc ttgtatatcc attttcggag
     2701 gttgggcttc ggaatcgttt tccgggacgc cggctggatg atcctccagc gcggggatct
     2761 catgctggag ttcttcgccc accccaactt gtttattgca gcttataatg gttacaaata
     2821 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     2881 tttgtccaaa ctcatcaatg tatcttatca tgtctgtata ccgtcgacct ctagctagag
     2941 cttggcgtaa tcatggtcat agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc
     3001 acacaacata cgagccggaa gcataaagtg taaagcctgg ggtgcctaat gagtgagcta
     3061 actcacatta attgcgttgc gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca
     3121 gctgcattaa tgaatcggcc aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc
     3181 cgcttcctcg ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc
     3241 tcactcaaag gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat
     3301 gtgagcaaaa ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt
     3361 ccataggctc cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg
     3421 aaacccgaca ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc
     3481 tcctgttccg accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt
     3541 ggcgctttct catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa
     3601 gctgggctgt gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta
     3661 tcgtcttgag tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa
     3721 caggattagc agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa
     3781 ctacggctac actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt
     3841 cggaaaaaga gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt
     3901 ttttgtttgc aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat
     3961 cttttctacg gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat
     4021 gagattatca aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc
     4081 aatctaaagt atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc
     4141 acctatctca gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta
     4201 gataactacg atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga
     4261 tccacgctca ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg
     4321 cagaagtggt cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc
     4381 tagagtaagt agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat
     4441 cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag
     4501 gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat
     4561 cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa
     4621 ttctcttact gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa
     4681 gtcattctga gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga
     4741 taataccgcg ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg
     4801 gcgaaaactc tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc
     4861 acccaactga tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg
     4921 aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact
     4981 cttccttttt caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat
     5041 atttgaatgt atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt
     5101 gccacctgac gtcgacggat cggg
//
