LOCUS       sequence-214283-        7273 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses a zinc finger specific to the human VEGFA locus, which is 
            linked to a KRAB transcriptional repression domain via a StaPL(TI) 
            module, in mammalian cells. [TI = telaprevir inhibited]..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7273)
  AUTHORS   Jacobs CL, Badiee RK, Lin MZ
  TITLE     StaPLs: versatile genetically encoded modules for engineering 
            drug-inducible proteins.
  JOURNAL   Nat Methods. 2018 Jul;15(7):523-526. doi: 10.1038/s41592-018-0041-z.
            Epub 2018 Jul 2.
  PUBMED    29967496
REFERENCE   2  (bases 1 to 7273)
  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"; doi: 
            "10.1038/s41592-018-0041-z"; journalName: "Nat Methods"; date: 
            "2018-07"; volume: "15"; issue: "7"; pages: "523-526"
COMMENT     Sequence Label: pCS6-ZF(VEGFA)-StaPL(TI)-tdRFP-KRAB
FEATURES             Location/Qualifiers
     source          1..7273
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     505..529
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     primer_bind     634..650
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     634..650
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     promoter        661..679
                     /label=SP6 promoter
                     /note="promoter for bacteriophage SP6 RNA polymerase"
     primer_bind     661..678
                     /label=SP6
                     /note="SP6 promoter, forward primer"
     protein_bind    680..704
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     regulatory      723..732
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1038..1064
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1791..3218
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPY
                     EGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVM
                     NFEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGV
                     LKGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYE
                     RSEGRHHLFLYGMDELYK"
     CDS             3267..3461
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the human zinc finger protein ZNF10 
                     (Margolin et al., 1994)"
                     /label=KRAB
                     /translation="RTLVTFKDVFVDFTREEWKLLDTAQQIVYRNVMLENYKNLVSLGY
                     QLTKPDVILRLEKGEEPWLV"
     CDS             3594..3614
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             3618..3638
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     protein_bind    complement(3677..3701)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     primer_bind     complement(3702..3721)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(3703..3721)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     primer_bind     complement(3732..3749)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(3732..3748)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3741..3763)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     polyA_signal    4153..4287
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4190..4209)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4244..4263
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      4413..4867
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4500..4519)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4709..4730
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     CDS             join(5154..5222,5223..6014)
                     /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     complement(5372..5391)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     protein_bind    6116..6149
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     rep_origin      6227..6815
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6716..6735
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     6969..6986
                     /label=L4440
                     /note="L4440 vector, forward primer"
     promoter        complement(7060..7164)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctcg tttagtgaac cgtcagatcg cctggagacg
      541 ccatccacgc tgttttgacc tccatagaag acaccgggac cgatccagcc tccggactct
      601 agcctaggcc gcgggacgga taacaatttc acacaggaaa cagctatgac cattaggcct
      661 atttaggtga cactatagaa caagtttgta caaaaaagca ggctggtacc ggtccggaat
      721 tcgccaccat gggcaaaaag aagcagcaca tctgccacat ccagggatgc ggcaaggtgt
      781 acggcaccac cagcaacctg aggaggcacc ttaggtggca caccggggag aggcccttca
      841 tgtgcacctg gagctactgc ggaaaaaggt tcaccaggag cagcaacttg cagaggcaca
      901 agaggaccca cacaggcgag aagaagttcg cctgccccga gtgccccaag aggtttatga
      961 ggagcgacca tctgagcagg cacatcaaga cccaccagaa caagaaaggc gggggcagtg
     1021 gcgggagcgg gcgcgcctat ccctacgatg tgcccgatta cgctggtgga tccggcgggt
     1081 cagatgagat ggaagagtgc tctcagcacg gcggatctgg aggttctaca ggctgcgtgg
     1141 tcatagtggg caggatcgtc ttgtccggat ccggcactag tgcgcccatc acggcgtacg
     1201 cccagcagac gagaggcctc ctagggtgta taatcaccag cctgactggc cgggacaaaa
     1261 accaagtgga gggtgaggtc cagatcgtgt caactgctac ccaaaccttg ctggcaacgt
     1321 gcatcaatgg ggtatgctgg acagtctacc acggggccgg aacgaggacc atcgcatcac
     1381 ccaagggtcc tgtcatccag atgtatacca atgtggacaa agaccttgtg ggctggcccg
     1441 ctcctcaagg ttcccgctca ttgacaccct gtacctgcgg ctcctcggac ctttacctgg
     1501 tcacgaggca cgccgatgtc attcccgtgc gccggcgagg tgatcgcagg ggtagcctgc
     1561 tttcgccccg gcccatttcc tacttgaaag gctcctcggg gggtccgctg ttgtgccccg
     1621 cgggacacgc cgtgggccta ttcagggccg cggtgtgcac ccgtggagtg gctaaagcgg
     1681 tgtactttat ccctgtggag aacctagaga caaccatgag atccccggtg ttcacggaca
     1741 actcctctcc accagcagtc accctgacgc acgggggttc aggcggtagt atggtgagca
     1801 agggcgagga ggtcatcaaa gagttcatgc gcttcaaggt gcgcatggag ggctccatga
     1861 acggccacga gttcgagatc gagggcgagg gcgagggccg cccctacgag ggcacccaga
     1921 ccgccaagct gaaggtgacc aagggcggcc ccctgccctt cgcctgggac atcctgtccc
     1981 cccagttcat gtacggctcc aaggcgtacg tgaagcaccc cgccgacatc cccgattaca
     2041 agaagctgtc cttccccgag ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg
     2101 gtctggtgac cgtgacccag gactcctccc tgcaggacgg cacgctgatc tacaaggtga
     2161 agatgcgcgg caccaacttc ccccccgacg gccccgtaat gcagaagaag accatgggct
     2221 gggaggcctc caccgagcgc ctgtaccccc gcgacggcgt gctgaagggc gagatccacc
     2281 aggccctgaa gctgaaggac ggcggccact acctggtgga gttcaagacc atctacatgg
     2341 ccaagaagcc cgtgcaactg cccggctact actacgtgga caccaagctg gacatcacct
     2401 cccacaacga ggactacacc atcgtggaac agtacgagcg ctccgagggc cgccaccacc
     2461 tgttcctggg gcatggcacc ggcagcaccg gcagcggcag ctccggcacc gcctcctccg
     2521 aggacaacaa catggccgtc atcaaagagt tcatgcgctt caaggtgcgc atggagggct
     2581 ccatgaacgg ccacgagttc gagatcgagg gcgagggcga gggccgcccc tacgagggca
     2641 cccagaccgc caagctgaag gtgaccaagg gcggccccct gcccttcgcc tgggacatcc
     2701 tgtcccccca gttcatgtac ggctccaagg cgtacgtgaa gcaccccgcc gacatccccg
     2761 attacaagaa gctgtccttc cccgagggct tcaagtggga gcgcgtgatg aacttcgagg
     2821 acggcggtct ggtgaccgtg acccaggact cctccctgca ggacggcacg ctgatctaca
     2881 aggtgaagat gcgcggcacc aacttccccc ccgacggccc cgtaatgcag aagaagacca
     2941 tgggctggga ggcctccacc gagcgcctgt acccccgcga cggcgtgctg aagggcgaga
     3001 tccaccaggc cctgaagctg aaggacggcg gccactacct ggtggagttc aagaccatct
     3061 acatggccaa gaagcccgtg caactgcccg gctactacta cgtggacacc aagctggaca
     3121 tcacctccca caacgaggac tacaccatcg tggaacagta cgagcgctcc gagggccgcc
     3181 accacctgtt cctgtacggc atggacgagc tgtacaaggg aggctccgga ggtagcatgg
     3241 acgccaagag cctgaccgcc tggagcagga cactggtgac cttcaaggac gtgttcgtgg
     3301 acttcacccg agaggagtgg aagctgctgg acaccgccca gcagatcgtg tacaggaacg
     3361 tgatgctgga aaactacaag aacctggtga gcctgggcta ccagctgacc aagcccgacg
     3421 tgatcctgag gctggagaag ggcgaggagc cctggttggt ggaaagggag atccaccagg
     3481 agacccaccc cgacagcgag accgcatttg agatcaagag cagcgtgggc ggatcaggcg
     3541 gctccgatga gatggaagag tgctctcagc acggcggctc cggcggctcc gaccctaaga
     3601 aaaaacggaa agttgatcca aaaaagaaga gaaaggtata gctagagtat ccctcgaggg
     3661 gcccaagctt acgcgtaccc agctttcttg tacaaagtgg tccctatagt gagtcgtatt
     3721 ataagctagg cactggccgt cgttttacaa cgtcgtgact gggaaaactg ctagcttggg
     3781 atctttgtga aggaacctta cttctgtggt gtgacataat tggacaaact acctacagag
     3841 atttaaagct ctaaggtaaa tataaaattt ttaagtgtat aatgtgttaa actagctgca
     3901 tatgcttgct gcttgagagt tttgcttact gagtatgatt tatgaaaata ttatacacag
     3961 gagctagtga ttctaattgt ttgtgtattt tagattcaca gtcccaaggc tcatttcagg
     4021 cccctcagtc ctcacagtct gttcatgatc ataatcagcc ataccacatt tgtagaggtt
     4081 ttacttgctt taaaaaacct cccacacctc cccctgaacc tgaaacataa aatgaatgca
     4141 attgttgttg ttaacttgtt tattgcagct tataatggtt acaaataaag caatagcatc
     4201 acaaatttca caaataaagc atttttttca ctgcattcta gttgtggttt gtccaaactc
     4261 atcaatgtat cttatcatgt ctggatcgat cctgcattaa tgaatcggcc aacgcgcggg
     4321 gagaggcggt ttgcgtattg gctggcgtaa tagcgaagag gcccgcaccg atcgcccttc
     4381 ccaacagttg cgcagcctga atggcgaatg ggacgcgccc tgtagcggcg cattaagcgc
     4441 ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt gccagcgccc tagcgcccgc
     4501 tcctttcgct ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct
     4561 aaatcggggg ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa
     4621 acttgattag ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc
     4681 tttgacgttg gagtccacgt tcttaatagt ggactcttgt tccaaactgg aacaacactc
     4741 aaccctatct cggtctattc ttttgattta taagggattt tgccgatttc ggcctattgg
     4801 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgctt
     4861 acaatttagg tggcactttt cggggaaatg tgcgcggaac ccctatttgt ttatttttct
     4921 aaatacattc aaatatgtat ccgctcatgc caggtcttgg actggtgaga acggcttgct
     4981 cggcagcttc gatgtgtgct ggagggagaa taaaggtcta agatgtgcga tagagggaag
     5041 tcgcattgaa ttatgtgctg tgtagggatc gctggtatca aatatgtgtg cccacccctg
     5101 gcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     5161 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     5221 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     5281 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     5341 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     5401 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     5461 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     5521 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     5581 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     5641 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     5701 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     5761 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     5821 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     5881 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     5941 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     6001 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     6061 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgccataac
     6121 ttcgtataat gtatgctata cgaagttatg gcatgaccaa aatcccttaa cgtgagtttt
     6181 cgttccactg agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt
     6241 ttctgcgcgt aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt
     6301 tgccggatca agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga
     6361 taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag
     6421 caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata
     6481 agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg
     6541 gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga
     6601 gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca
     6661 ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa
     6721 acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt
     6781 tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac
     6841 ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta tcccctgatt
     6901 ctgtggataa ccgtattacc gcctttgagt gagctgatac cgctcgccgc agccgaacga
     6961 ccgagcgcag cgagtcagtg agcgaggaag cggaagagcg cccaatacgc aaaccgcctc
     7021 tccccgcgcg ttggccgatt cattaatgca gagcttgcaa ttcgcgcttt ttcaatatta
     7081 ttgaagcatt tatcagggtt attgtctcat gagcggatac atatttgaat gtatttagaa
     7141 aaataaacaa ataggggttc cgcgcacatt tccccgaaaa gtgccacctg acgtctaaga
     7201 aaccattatt atcatgacat taacctataa aaataggcgt attacgaggc cctttcacgc
     7261 attagatgca tgt
//
