LOCUS       sequence-318786-        5481 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Inducible expression of 3xFLAG-NLS-DsDed-ZF10 under the ZF6x6-C 
            promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5481)
  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 5481)
  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: pJM575 ZF6x6-C 3xFLAG-NLS-DsDed-ZF10 in TUPV2
FEATURES             Location/Qualifiers
     source          1..5481
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     regulatory      213..222
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             222..287
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     CDS             294..314
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             357..1031
                     /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(537..557)
                     /label=DsRed1-N
                     /note="DsRed1, reverse primer"
     primer_bind     949..972
                     /label=DsRed1-C
                     /note="DsRed1, forward primer"
     primer_bind     complement(1388..1405)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    1394..1618
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      2387..2815
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2474..2493)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2684..2705
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        2838..3034
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      2885..3020
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     2947..2966
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     polyA_signal    3154..3275
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3191..3210)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3245..3264
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(3324..3340)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3324..3340)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3337..3359)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3348..3364
                     /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(3372..3378,3379..3396,3397..3402))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3417..3438
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3555..3572)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3726..4314)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3806..3825)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4485..5276,5277..5345))
                     /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     5108..5127
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5346..5450)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatcgaag acgggcaaaa ccgatgttga cggactaatc ctgacaagat
       61 ctgcgaggtg taggggacgc gagtgtaggg gtgagctgtg tagggggact aagtgtaggg
      121 gacgcgagtg taggggtgag ctgtgtaggg gaacgctctc tagagggtat ataatggggg
      181 ccactagtct actaccagag ctcatcgcta gcgccaccat ggattataag gatcacgatg
      241 gcgactacaa ggaccacgat attgactaca aagacgatga cgataaaatg gctcctaaga
      301 aaaagcgcaa agtcggtatc catggcgttc ccggatcagg ctccggaagt ggctccatgg
      361 atagcactga gaacgtcatc aagcccttca tgcgcttcaa ggtgcacatg gagggctccg
      421 tgaacggcca cgagttcgag atcgagggcg agggcgaggg caagccctac gagggcaccc
      481 agaccgccaa gctccaggtg accaagggcg gccccctgcc cttcgcctgg gacatcctgt
      541 ccccccagtt ccagtacggc tccaaggtgt acgtgaagca ccccgccgac atccccgact
      601 acaagaagct gtccttcccc gagggcttca agtgggagaa ggtgatgaac ttcgaggacg
      661 gcggcgtggt gaccgtgacc caggactcct ccctccagga cggcaccttc atctaccacg
      721 tgaagttcat cggcgtgaac ttcccctccg acggccccgt aatgcagaag aaaactctgg
      781 gctgggagcc ctccaccgag cgcctgtacc cccgcgacgg cgtgctgaag ggcgagatcc
      841 acaaggcgct gaagctgaag ggcggcggcc actacctggt ggagttcaag tcaatctaca
      901 tggccaagaa gcccgtgaag ctgcccggct actactacgt ggactccaag ctggacatca
      961 cctcccacaa cgaggactac accgtggtgg agcagtacga gcgcgccgag gcccgccacc
     1021 acctgttcca gtctggttct ggttctggtt caggatccgg taccggtgga gctagacccg
     1081 gggagcgccc cttccagtgt cgcatttgca tgcggaactt ttcgactaat caaaaattgg
     1141 aagttcatac ccgtactcat accggtgaaa aaccgtttca gtgtcggatc tgtatgcgaa
     1201 atttctccgt tagacataat ttgcaaagac atctacgtac gcacaccggc gagaagccat
     1261 tccaatgccg aatatgcatg cgcaacttca gtcaacatcc aaatttgact agacacctaa
     1321 aaacccacct gagaggatct taagcggccg ctcgagtcta gagggcccgt ttaaacccgc
     1381 tgatcagcct cgactgtgcc ttctagttgc cagccatctg ttgtttgccc ctcccccgtg
     1441 ccttccttga ccctggaagg tgccactccc actgtccttt cctaataaaa tgaggaaatt
     1501 gcatcgcatt gtctgagtag gtgtcattct attctggggg gtggggtggg gcaggacagc
     1561 aagggggagg attgggaagt caatagcagg catgctgggg atgcggtggg ctctatggct
     1621 tctgaggcag gtcgaagaac tatttcctgg ctgttacgcg agcgcatcgc gataagctct
     1681 agagggacag ccccccccca aagcccccag ggatgtaatt acgtccctcc cccgctaggg
     1741 ggcagcagcg agccgcccgg ggctccgctc cggtccggcg ctccccccgc atccccgagc
     1801 cggcagcgtg cggggacagc ccgggcacgg ggaaggtggc acgggatcgc tttcctctga
     1861 acgcttctcg ctgctctttg agcctgcaga cacctggggg gatacgggga aaaagcttta
     1921 ggctgaaaga gagatttaga atgacagaat catagaacgg cctgcgtacg gatggggctc
     1981 tagctctaga gcttgatatc gaattcctgc agccccgcta gagggacagc ccccccccaa
     2041 agcccccagg gatgtaatta cgtccctccc ccgctagggg gcagcagcga gccgcccggg
     2101 gctccgctcc ggtccggcgc tccccccgca tccccgagcc ggcagcgtgc ggggacagcc
     2161 cgggcacggg gaaggtggca cgggatcgct ttcctctgaa cgcttctcgc tgctctttga
     2221 gcctgcagac acctgggggg atacggggaa aaagctttag gctgaaagag agatttagaa
     2281 tgacagaatc atagaacggc ctgggcgcgc ctggtggcga acgatcaaca gacgggagag
     2341 gactaaagtc ttcgctgaca gcgaccggac tctagggggt atccccacgc gccctgtagc
     2401 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     2461 gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     2521 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     2581 ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc gccctgatag
     2641 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     2701 actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg gattttgccg
     2761 atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattaattc
     2821 tgtggaatgt gtgtcagtta ggctcaatta gtcagcaacc atagtcccgc ccctaactcc
     2881 gcccatcccg cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat
     2941 tttttttatt tatgcagagg ccgaggccgc ctctgcctct gagctattcc agaagtagtg
     3001 aggaggcttt tttggaggcc taggcttttg caaaaagctc ccgggagctt gtatatccat
     3061 tttcggaggt tgggcttcgg aatcgttttc cgggacgccg gctggatgat cctccagcgc
     3121 ggggatctca tgctggagtt cttcgcccac cccaacttgt ttattgcagc ttataatggt
     3181 tacaaataaa gcaatagcat cacaaatttc acaaataaag catttttttc actgcattct
     3241 agttgtggtt tgtccaaact catcaatgta tcttatcatg tctgtatacc gtcgacctct
     3301 agctagagct tggcgtaatc atggtcatag ctgtttcctg tgtgaaattg ttatccgctc
     3361 acaattccac acaacatacg agccggaagc ataaagtgta aagcctgggg tgcctaatga
     3421 gtgagctaac tcacattaat tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg
     3481 tcgtgccagc tgcattaatg aatcggccaa cgcgcgggga gaggcggttt gcgtattggg
     3541 cgctcttccg cttcctcgct cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg
     3601 gtatcagctc actcaaaggc ggtaatacgg ttatccacag aatcagggga taacgcagga
     3661 aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg
     3721 gcgtttttcc ataggctccg cccccctgac gagcatcaca aaaatcgacg ctcaagtcag
     3781 aggtggcgaa acccgacagg actataaaga taccaggcgt ttccccctgg aagctccctc
     3841 gtgcgctctc ctgttccgac cctgccgctt accggatacc tgtccgcctt tctcccttcg
     3901 ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt
     3961 cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc
     4021 ggtaactatc gtcttgagtc caacccggta agacacgact tatcgccact ggcagcagcc
     4081 actggtaaca ggattagcag agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg
     4141 tggcctaact acggctacac tagaagaaca gtatttggta tctgcgctct gctgaagcca
     4201 gttaccttcg gaaaaagagt tggtagctct tgatccggca aacaaaccac cgctggtagc
     4261 ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat
     4321 cctttgatct tttctacggg gtctgacgct cagtggaacg aaaactcacg ttaagggatt
     4381 ttggtcatga gattatcaaa aaggatcttc acctagatcc ttttaaatta aaaatgaagt
     4441 tttaaatcaa tctaaagtat atatgagtaa acttggtctg acagttacca atgcttaatc
     4501 agtgaggcac ctatctcagc gatctgtcta tttcgttcat ccatagttgc ctgactcccc
     4561 gtcgtgtaga taactacgat acgggagggc ttaccatctg gccccagtgc tgcaatgata
     4621 ccgcgagatc cacgctcacc ggctccagat ttatcagcaa taaaccagcc agccggaagg
     4681 gccgagcgca gaagtggtcc tgcaacttta tccgcctcca tccagtctat taattgttgc
     4741 cgggaagcta gagtaagtag ttcgccagtt aatagtttgc gcaacgttgt tgccattgct
     4801 acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt cattcagctc cggttcccaa
     4861 cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa aagcggttag ctccttcggt
     4921 cctccgatcg ttgtcagaag taagttggcc gcagtgttat cactcatggt tatggcagca
     4981 ctgcataatt ctcttactgt catgccatcc gtaagatgct tttctgtgac tggtgagtac
     5041 tcaaccaagt cattctgaga atagtgtatg cggcgaccga gttgctcttg cccggcgtca
     5101 atacgggata ataccgcgcc acatagcaga actttaaaag tgctcatcat tggaaaacgt
     5161 tcttcggggc gaaaactctc aaggatctta ccgctgttga gatccagttc gatgtaaccc
     5221 actcgtgcac ccaactgatc ttcagcatct tttactttca ccagcgtttc tgggtgagca
     5281 aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg cgacacggaa atgttgaata
     5341 ctcatactct tcctttttca atattattga agcatttatc agggttattg tctcatgagc
     5401 ggatacatat ttgaatgtat ttagaaaaat aaacaaatag gggttccgcg cacatttccc
     5461 cgaaaagtgc cacctgacgt c
//
