LOCUS       sequence-317922-        5760 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of 3xFLAG-NLS-DsDed-intC-ZF1-NLS-HA under the 
            ZF6x6-C promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5760)
  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 5760)
  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: pJM573 ZF6x6-C 3xFLAG-NLS-DsDed-intC-ZF1-NLS-H...
FEATURES             Location/Qualifiers
     source          1..5760
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      258..267
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             267..332
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             339..359
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             402..1076
                     /codon_start=1
                     /product="noncytotoxic tetrameric variant of DsRed 
                     fluorescent protein (Strack et al., 2008)"
                     /label=DsRed-Express2
                     /note="mammalian codon-optimized"
                     /translation="MDSTENVIKPFMRFKVHMEGSVNGHEFEIEGEGEGKPYEGTQTAK
                     LQVTKGGPLPFAWDILSPQFQYGSKVYVKHPADIPDYKKLSFPEGFKWEKVMNFEDGGV
                     VTVTQDSSLQDGTFIYHVKFIGVNFPSDGPVMQKKTLGWEPSTERLYPRDGVLKGEIHK
                     ALKLKGGGHYLVEFKSIYMAKKPVKLPGYYYVDSKLDITSHNEDYTVVEQYERAEARHH
                     LFQ"
     primer_bind     complement(582..602)
                     /label=DsRed1-N
                     /note="DsRed1, reverse primer"
     primer_bind     994..1017
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     CDS             1605..1625
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             1638..1664
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     primer_bind     complement(1712..1729)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1718..1942
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2711..3139
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2798..2817)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3008..3029
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        3162..3358
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      3209..3344
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3271..3290
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    3478..3599
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3515..3534)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3569..3588
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3648..3664)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3648..3664)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3661..3683)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3672..3688
                     /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(3696..3702,3703..3720,3721..3726))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3741..3762
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3879..3896)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4050..4638)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4130..4149)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4809..5600,5601..5669))
                     /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     5432..5451
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(join(5670..5760,1..14))
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtcgacgg atcgggagat
       61 cgaagacggg caaaaccgat gttgacggac taatcctgac aagatctgcg aggtgtaggg
      121 gacgcgagtg taggggtgag ctgtgtaggg ggactaagtg taggggacgc gagtgtaggg
      181 gtgagctgtg taggggaacg ctctctagag ggtatataat gggggccact agtctactac
      241 cagagctcat cgctagcgcc accatggatt ataaggatca cgatggcgac tacaaggacc
      301 acgatattga ctacaaagac gatgacgata aaatggctcc taagaaaaag cgcaaagtcg
      361 gtatccatgg cgttcccgga tcaggctccg gaagtggctc catggatagc actgagaacg
      421 tcatcaagcc cttcatgcgc ttcaaggtgc acatggaggg ctccgtgaac ggccacgagt
      481 tcgagatcga gggcgagggc gagggcaagc cctacgaggg cacccagacc gccaagctcc
      541 aggtgaccaa gggcggcccc ctgcccttcg cctgggacat cctgtccccc cagttccagt
      601 acggctccaa ggtgtacgtg aagcaccccg ccgacatccc cgactacaag aagctgtcct
      661 tccccgaggg cttcaagtgg gagaaggtga tgaacttcga ggacggcggc gtggtgaccg
      721 tgacccagga ctcctccctc caggacggca ccttcatcta ccacgtgaag ttcatcggcg
      781 tgaacttccc ctccgacggc cccgtaatgc agaagaaaac tctgggctgg gagccctcca
      841 ccgagcgcct gtacccccgc gacggcgtgc tgaagggcga gatccacaag gcgctgaagc
      901 tgaagggcgg cggccactac ctggtggagt tcaagtcaat ctacatggcc aagaagcccg
      961 tgaagctgcc cggctactac tacgtggact ccaagctgga catcacctcc cacaacgagg
     1021 actacaccgt ggtggagcag tacgagcgcg ccgaggcccg ccaccacctg ttccagtctg
     1081 gttctggttc tggttcagga tccggtggtg gcgggtctgg tggaggaggt agtggtggag
     1141 gtggctctgc caccatgatg ctgaagaaga tcctgaagat cgaggagctg gacgagagag
     1201 agctgatcga catcgaggtg agcggcaacc acctgttcta cgccaacgac atcctgaccc
     1261 acaacagcaa caggggtggt ggcgggtctg gtggaggagg tagtggtgga ggtggctctg
     1321 gtaccgctag acccggggag cgccccttcc agtgtcgcat ttgcatgcgg aacttttcgc
     1381 gccaggacag gcttgacagg catacccgta ctcataccgg tgaaaaaccg tttcagtgtc
     1441 ggatctgtat gcgaaatttc tcccagaagg agcacttggc ggggcatcta cgtacgcaca
     1501 ccggcgagaa gccattccaa tgccgaatat gcatgcgcaa cttcagtcgc cgcgacaacc
     1561 tgaaccggca cctaaaaacc cacctgagag aattcggctc tggccctaag aagaagcgca
     1621 aagttggctc tggcatgtac ccatacgacg taccagatta cgcttaggcg gccgctcgag
     1681 tctagagggc ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca
     1741 tctgttgttt gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc
     1801 ctttcctaat aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg
     1861 gggggtgggg tggggcagga cagcaagggg gaggattggg aagtcaatag caggcatgct
     1921 ggggatgcgg tgggctctat ggcttctgag gcaggtcgaa gaactatttc ctggctgtta
     1981 cgcgagcgca tcgcgataag ctctagaggg acagcccccc cccaaagccc ccagggatgt
     2041 aattacgtcc ctcccccgct agggggcagc agcgagccgc ccggggctcc gctccggtcc
     2101 ggcgctcccc ccgcatcccc gagccggcag cgtgcgggga cagcccgggc acggggaagg
     2161 tggcacggga tcgctttcct ctgaacgctt ctcgctgctc tttgagcctg cagacacctg
     2221 gggggatacg gggaaaaagc tttaggctga aagagagatt tagaatgaca gaatcataga
     2281 acggcctgcg tacggatggg gctctagctc tagagcttga tatcgaattc ctgcagcccc
     2341 gctagaggga cagccccccc ccaaagcccc cagggatgta attacgtccc tcccccgcta
     2401 gggggcagca gcgagccgcc cggggctccg ctccggtccg gcgctccccc cgcatccccg
     2461 agccggcagc gtgcggggac agcccgggca cggggaaggt ggcacgggat cgctttcctc
     2521 tgaacgcttc tcgctgctct ttgagcctgc agacacctgg ggggatacgg ggaaaaagct
     2581 ttaggctgaa agagagattt agaatgacag aatcatagaa cggcctgggc gcgcctggtg
     2641 gcgaacgatc aacagacggg agaggactaa agtcttcgct gacagcgacc ggactctagg
     2701 gggtatcccc acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     2761 agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc
     2821 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     2881 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca
     2941 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     3001 tttaatagtg gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct
     3061 tttgatttat aagggatttt gccgatttcg gcctattggt taaaaaatga gctgatttaa
     3121 caaaaattta acgcgaatta attctgtgga atgtgtgtca gttaggctca attagtcagc
     3181 aaccatagtc ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca
     3241 ttctccgccc catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc
     3301 ctctgagcta ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa
     3361 gctcccggga gcttgtatat ccattttcgg aggttgggct tcggaatcgt tttccgggac
     3421 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     3481 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     3541 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     3601 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
     3661 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     3721 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     3781 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     3841 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     3901 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     3961 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     4021 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     4081 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     4141 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     4201 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     4261 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     4321 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     4381 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     4441 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     4501 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     4561 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     4621 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     4681 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     4741 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     4801 tctgacagtt accaatgctt aatcagtgag gcacctatct cagcgatctg tctatttcgt
     4861 tcatccatag ttgcctgact ccccgtcgtg tagataacta cgatacggga gggcttacca
     4921 tctggcccca gtgctgcaat gataccgcga gatccacgct caccggctcc agatttatca
     4981 gcaataaacc agccagccgg aagggccgag cgcagaagtg gtcctgcaac tttatccgcc
     5041 tccatccagt ctattaattg ttgccgggaa gctagagtaa gtagttcgcc agttaatagt
     5101 ttgcgcaacg ttgttgccat tgctacaggc atcgtggtgt cacgctcgtc gtttggtatg
     5161 gcttcattca gctccggttc ccaacgatca aggcgagtta catgatcccc catgttgtgc
     5221 aaaaaagcgg ttagctcctt cggtcctccg atcgttgtca gaagtaagtt ggccgcagtg
     5281 ttatcactca tggttatggc agcactgcat aattctctta ctgtcatgcc atccgtaaga
     5341 tgcttttctg tgactggtga gtactcaacc aagtcattct gagaatagtg tatgcggcga
     5401 ccgagttgct cttgcccggc gtcaatacgg gataataccg cgccacatag cagaacttta
     5461 aaagtgctca tcattggaaa acgttcttcg gggcgaaaac tctcaaggat cttaccgctg
     5521 ttgagatcca gttcgatgta acccactcgt gcacccaact gatcttcagc atcttttact
     5581 ttcaccagcg tttctgggtg agcaaaaaca ggaaggcaaa atgccgcaaa aaagggaata
     5641 agggcgacac ggaaatgttg aatactcata ctcttccttt ttcaatatta ttgaagcatt
     5701 tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa aaataaacaa
//
