LOCUS       sequence-375608-        9586 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  PiggyBAC vector expressing a FOS promoter driven ""KGV"" synthetic 
            transcription factor and a CMV promoter driven BFP marker..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 9586)
  AUTHORS   Ravindran PT, McFann S, Thornton RH, Toettcher JE
  TITLE     A synthetic gene circuit for imaging-free detection of signaling 
            pulses.
  JOURNAL   Cell Syst. 2022 Feb 16;13(2):131-142.e13. doi: 
            10.1016/j.cels.2021.10.002. Epub 2021 Nov 4.
  PUBMED    34739875
REFERENCE   2  (bases 1 to 9586)
  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: 
            "10.1016/j.cels.2021.10.002"; journalName: "Cell Syst"; date: 
            "2022-02-16- 16"; volume: "13"; issue: "2"; pages: "131-142.e13"
COMMENT     Sequence Label: pPBJ pFOS-KTR-Gal4-PEST-tub3' pCMV-TagBFP
FEATURES             Location/Qualifiers
     source          1..9586
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   23..57
                     /label=piggyBac left (5') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     CDS             complement(2869..2880)
                     /codon_start=1
                     /product="WELQut protease recognition and cleavage site"
                     /label=WELQut site
                     /translation="WELQ"
     CDS             3229..3669
                     /codon_start=1
                     /gene="Saccharomyces cerevisiae GAL4 (truncated)"
                     /product="DNA binding domain of the GAL4 transcriptional 
                     activator"
                     /label=GAL4 DNA binding domain
                     /translation="MKLLSSIEQACDICRLKKLKCSKEKPKCAKCLKNNWECRYSPKTK
                     RSPLTRAHLTEVESRLERLEQLFLLIFPREDLDMILKMDSLQDIKALLTGLFVQDNVNK
                     DAVTDRLASVETDMPLTLRQHRISATSSSEESSNKGQRQLTVS"
     CDS             3706..3855
                     /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(5690..5710)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5695..6024
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5875..6010
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6034..6732
                     /codon_start=1
                     /product="monomeric blue fluorescent protein"
                     /label=TagBFP
                     /note="mammalian codon-optimized"
                     /translation="MSELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTMRIKV
                     VEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLTA
                     TQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRNDMALK
                     LVGGSHLIANIKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAVARYC
                     DLPSKLGHKLN"
     repeat_region   7009..7071
                     /label=piggyBac right (3') inverted repeat
                     /note="piggyBac transposon-specific inverted terminal 
                     repeat sequence (ITR)"
     primer_bind     complement(7183..7202)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     7302..7324
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(7362..7380)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        7448..7552
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(7553..7621,7622..8413)
                     /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(7771..7790)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      8584..9172
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     9073..9092
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     9326..9343
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    9460..9481
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(9496..9501,9502..9519,9520..9526)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    9534..9550
                     /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)."
     primer_bind     9539..9561
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     9558..9574
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     9558..9574
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 ggggcgcgcc attctagatt aaccctagaa agatagtctg cgtaaaattg acgcatgcat
       61 tcttgaaata ttgctctctc tttctaaata gcgcgaatcc gtcgctgtgc atttaggaca
      121 tctcagtcgc cgcttggagc tcccgtgagg cgtgcttgtc aatgcggtaa gtgtcactga
      181 ttttgaacta taacgaccgc gtgagtcaaa atgacgcatg attatctttt acgtgacttt
      241 taagatttaa ctcatacgat aattatattg ttatttcatg ttctacttac gtgataactt
      301 attatatata tattttcttg ttatagatat caactagaat gctagcatgg gcccagatct
      361 ccgaggggac ccgacaggcc cgcagggaca gcagagatcc agtttggatc gataagcttg
      421 atatcgaatt gacctctctt tttttactgc caaagtaagt gatgtatgag ataaatcctg
      481 gatataaaat cttgacacat ctttgaggtg ttggccagag ctgcttcaaa cctgaaaagg
      541 aaataaatca aatctctgca gtttttctct aagagtccct gagacgcagt tatgagtgga
      601 aggcaaggcc agacagaccc aggaactcaa aggttgacct actttgtcga aattaacaag
      661 agggaagacg tctgcataat ttttctgtct ctccctttca cagtaaagaa cggtgggagc
      721 acatgacaga cgagctgcag gccacgaaat cccttcagga ttgagcagct acacgaagcc
      781 tgtgtttcta tatcgtgttg ttctctttcc ctcccccctc ttccgagtga gaaaaaaaag
      841 gggggcctaa attcccacca acataaacta atgacataca atgatgaaat tctgttttca
      901 cctctgcctg tgacagggaa tgcaaaaata gcaagtggcc tatttccacg aatccccgcc
      961 tccctgccct ccccgctccc gctgggtttg gatctctaag aatggaggct aacgcagagg
     1021 caggagccag ccgggatcag ccccccgaac ccgagggtga tgtcagtcgg cgggagcgga
     1081 cgcagcctgg ccagggaggc agaggttcct agggagcagg gagggcgggt ggcctggatg
     1141 ggcggaccgt gcgcccggcc gctccgtcgc ctggagacgg ccctggcggc gctgtgttgc
     1201 tgtaaacagc cgcttcgctg ttactatcta tagaaagatc gcctagctcc cgggtgcggc
     1261 ggcccgaggc aggtttgtgg ttctgcaccc cttgcgcccc taaagatcca tccgctgtcc
     1321 taatcctctc tagtccgtct tggaatgccc accgtttggc ttgacccttg ggaccctcag
     1381 aatggagacg agggggacac atgagttctg cgaggatctg cggtttccta tcccagaggt
     1441 gaccggccca gtcagtctaa cccggcttgt cctctgcgga aggacaggag gccgagggca
     1501 agtaggggtg tgtttgttct acactgaagc acctgacctc ttcaaagttc catcttccaa
     1561 gactcaaagc tgttctcagg tcccagacgc caaaatctcg gcacagctgg gaacctttct
     1621 tcccgtcccc tctgcgcccc cacccccctt cccaagtccg atctggaaaa tcacccgctg
     1681 caggcgggtt ccttgtaagc gcagtttcca ggctgcacgt attcagaccc ccatctcccc
     1741 agcaccgact tgctttctcc tccccccccc cccgagctca cctcactttg taattctgag
     1801 ctccccccct gccccgactc gccctctggt ctcagctcaa aactaaacat acgacccctt
     1861 caggcatact tgtagggtgg ttttgcacaa tgtttatccg tcagtgtcaa cggggactgt
     1921 cgccttgata gctctaagtg gctaagggtc ggggagtagg tgctgccgtc ctttaaaaca
     1981 cgaatttatg aatgaaccca gtactgtagt taaatcaggt tattgtacac ttatttacaa
     2041 tccttcactt gctgcttcca acctcagtcc taaagtttct ccaggcaagg agctggagag
     2101 aggggctgag aagctgaccc cccctttttc ttctctgcac tgatttggga tggggggctg
     2161 atgtgggcaa gctttccttt aggaacagag gcttcgagcc tttaaggctg cgtacttgct
     2221 tctcctaata ccagagactc aaaaaaaaaa aaaaagttcc agattgctgg acaatgaccc
     2281 gggtctcatc ccttgaccct gggaaccggg tccacattga atcaggtgcg aatgttcgct
     2341 cgccttctct gcctttcccg cctcccctcc cccggccgcg gccccggttc cccccctgcg
     2401 ctgcaccctc agagttggct gcagccggcg agctgttccc gtcaatccct ccctccttta
     2461 cacaggatgt ccatattagg acatctgcgt cagcaggttt ccacggccgg tccctgttgt
     2521 tctggggggg ggaccatctc cgaaatccta cacgcggaag gtctaggaga ccccctaaga
     2581 tcccaaatgt gaacactcat aggtgaaaga tgtatgccaa gacgggggtt gaaagcctgg
     2641 ggcgtagagt tgacgacaga gcgcccgcag agggccttgg ggcgcgcttc cccccccttc
     2701 cagttccgcc cagtgacgta ggaagtccat ccattcacag cgcttctata aaggcgccag
     2761 ctgaggcgcc tactactcca accgcgactg cagcgagcaa ctgagaagac tggatagagc
     2821 cggcggttcc gcgaacgagc agtgaccgcg ctcccaccca gctctgctct gcagctccca
     2881 ccagtgtcta cccctggacc ccttgccggg ctttccccaa acttcgacat gaagggccga
     2941 aagcctcggg acctggaact tccacttagc ccaagcctgc tgggtggcca gggacctgaa
     3001 cggactccag gatcaggaac aagctctggt cttcaggcac cggggccagc gctttcgccc
     3061 agcaagcgta gcggtctcga ggacccagcg acgccatcga agaagccacg taccccatcg
     3121 gtgtcgtcaa ggctggagcg cctgaccctc cagtccagct ttcaattccc atccactagt
     3181 acgcgtcaag tggagcaagg caggtggaca gtggatccgg tcgccaccat gaagctgctg
     3241 agcagcatcg agcaggcctg tgacatctgc cggctgaaga aactgaagtg cagcaaagaa
     3301 aagcccaagt gcgccaagtg cctgaagaac aactgggagt gccggtacag ccccaagacc
     3361 aagagaagcc ccctgaccag agcccacctg accgaggtgg aaagccggct ggaaagactg
     3421 gaacagctgt ttctgctgat cttcccacgc gaggacctgg acatgatcct gaagatggac
     3481 agcctgcagg acatcaaggc cctgctgacc ggcctgttcg tgcaggacaa cgtgaacaag
     3541 gacgccgtga ccgacagact ggccagcgtg gaaaccgaca tgcccctgac cctgcggcag
     3601 cacagaatca gcgccaccag cagcagcgag gaaagcagca acaagggcca gcggcagctg
     3661 acagtgtctg ctgctgcagg cggaagcgga ggctctggcg gatctgatgc cctggacgac
     3721 ttcgacctgg atatgctggg cagcgacgcc ctggatgatt ttgatctgga catgctggga
     3781 tctgacgctc tggacgattt cgatctcgac atgttgggat cagatgcact ggatgacttt
     3841 gacctggaca tgctcggatc aaagcttagc catggcttcc cgccggaggt ggaggagcag
     3901 gatgatggca cgctgcccat gtcttgtgcc caggagagcg ggatggaccg tcaccctgca
     3961 gcctgtgctt ctgctaggat caatgtgtga taagccgcga attcgccctt cgcgggatcc
     4021 tacggtactt attgccaaga aagcacgagc atcagccgtg cctccaggtc gaatcttcaa
     4081 acgacgacga tcacgcgtcg ctccagtatt ccagggttca tcagatccta cggtacttat
     4141 tgccaagaaa gcacgagcat cagccgtgcc tccaggtcga atcttcaaac gacgacgatc
     4201 acgcgtcgct ccagtattcc agggttcatc agatcctacg gtacttattg ccaagaaagc
     4261 acgagcatca gccgtgcctc caggtcgaat cttcaaacga cgacgatcac gcgtcgctcc
     4321 agtattccag ggttcatcag atcctacggt acttattgcc aagaaagcac gagcatcagc
     4381 cgtgcctcca ggtcgaatct tcaaacgacg acgatcacgc gtcgctccag tattccaggg
     4441 ttcatcagat ctcgcgaagg gcgaattcgt ttccatggcg cctgatgcgg tagcggccgc
     4501 ttcttaaagc ttttactttg agacatcatg gaaaacttaa gaggtacaac atggagaaga
     4561 catgatcaca gaatggaaac agcacagaag catcagtgcc tgcaactaat actggagcag
     4621 tttgacgaca caggggctca aggaatggac ttagtattcc tctccttctt cctccccctc
     4681 gccttccaca gaatccacac caacctcctc ataatccttc tctagggcag ccatgtcctc
     4741 acaggcctca gagaactcac cctcctccat gccctcaccc acataccagt gcacaaaggc
     4801 acgcttgaca tacatcagat caaacttgtg atctaggcga gcccaggcct cagcaatggc
     4861 tgtggtgttg ctcagcatgc acacagctct ctgcaccttg gccaggtcac caccgggtac
     4921 cacagtggga ggctggtagt taatgccaac cttgaagcca gtggggcacc agtctacaaa
     4981 ctggatgctg cgcttggtct tgatggtggc gatggcagca ttgacatctt tgggaactac
     5041 atcaccacgg tacagcaggc agcaagccat gtatttacca tggcgagggt cacatttcac
     5101 catctggttg gctggctcaa agcaggcatt ggtgatctct gctacagaaa gctgctcatg
     5161 gtaggctttc tcagcagaga tgacaggggc ataagtggcc agagggaagt ggatgcgagg
     5221 atagggtacc aagttggtct ggaattctgt cagatcaaca ttcagggccc atcaaatctg
     5281 agggaagcag tgatggaaga cacaatctgg ctaataaggc ggttaaggtt ggtgtaggtt
     5341 ggacgctcaa tgtcgaggtt tctacgacag atatcataga tggcctcatt gtctaccatg
     5401 aaggcacaat cagagtgctc cagggtggtg tgggtggtga ggatggaatt gtagggctca
     5461 accacagcag tggaaacctg gggggctggt taaatggaga actccagctt ggacttcttt
     5521 ccgtaatcca cagagagccg ctccatcagc agggaggtga agccagagcc agttcccccg
     5581 ccaaagctgt ggaaaaccaa gaagccctgg agacctgtgc actggtcagc tcgaccaagg
     5641 cagctgtaga tcttagccat ctgctttttg cttgtactgt cgaccctgtg gaatgtgtgt
     5701 cagttagggt gtggaaagtc cccaggctcc ccagcaggca gaagtatgca aagcatgcat
     5761 ctcaattagt cagcaaccag gtgtggaaag tccccaggct ccccagcagg cagaagtatg
     5821 caaagcatgc atctcaatta gtcagcaacc atagtcccgc ccctaactcc gcccatcccg
     5881 cccctaactc cgcccagttc cgcccattct ccgccccatg gctgactaat tttttttatt
     5941 tatgcagagg ccgaggccgc ctcggcctct gagctattcc agaagtagtg aggaggcttt
     6001 tttggaggcc taggcttttg caaaaagctt accatgagcg agctgattaa ggagaacatg
     6061 cacatgaagc tgtacatgga gggcaccgtg gacaaccatc acttcaagtg cacatccgag
     6121 ggcgaaggca agccctacga gggcacccag accatgagaa tcaaggtggt cgagggcggc
     6181 cctctcccct tcgccttcga catcctggct actagcttcc tctacggcag caagaccttc
     6241 atcaaccaca cccagggcat ccccgacttc ttcaagcagt ccttccctga gggcttcaca
     6301 tgggagagag tcaccacata cgaagacggg ggcgtgctga ccgctaccca ggacaccagc
     6361 ctccaggacg gctgcctcat ctacaacgtc aagatcagag gggtgaactt cacatccaac
     6421 ggccctgtga tgcagaagaa aacactcggc tgggaggcct tcaccgagac gctgtacccc
     6481 gctgacggcg gcctggaagg cagaaacgac atggccctga agctcgtggg cgggagccat
     6541 ctgatcgcaa acatcaagac cacatataga tccaagaaac ccgctaagaa cctcaagatg
     6601 cctggcgtct actatgtgga ctacagactg gaaagaatca aggaggccaa caacgagacc
     6661 tacgtcgagc agcacgaggt ggcagtggcc agatactgcg acctccctag caaactgggg
     6721 cacaagctta atgcggccgc aactcccacc tgcaacatgc gtgactaacg cccgccccac
     6781 gacccgcagc gcccgaccga aaggagcgca cgaccccatg catcgatact agttaaaagt
     6841 tttgttactt tatagaagaa attttgagtt tttgtttttt tttaataaat aaataaacat
     6901 aaataaattg tttgttgaat ttattattag tatgtaagtg taaatataat aaaacttaat
     6961 atctattcaa attaataaat aaacctcgat atacagaccg ataaaacaca tgcgtcaatt
     7021 ttacgcatga ttatctttaa cgtacgtcac aatatgatta tctttctagg gttaatctag
     7081 tatacgcgta tgcggccgct taattaatcc atggcgcctg atgcggtatt ttctccttac
     7141 gcatctgtgc ggtatttcac accgcatatg gtgcactctc agtacaatct gctctgatgc
     7201 cgcatagtta agccagcccc gacacccgcc aacacccgct gacgcgccct gacgggcttg
     7261 tctgctcccg gcatccgctt acagacaagc tgtgaccgtc tccgggagct gcatgtgtca
     7321 gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt
     7381 tttataggtt aatgtcatga taataatggt ttcttagacg tcaggtggca cttttcgggg
     7441 aaatgtgcgc ggaaccccta tttgtttatt tttctaaata cattcaaata tgtatccgct
     7501 catgagacaa taaccctgat aaatgcttca ataatattga aaaaggaaga gtatgagtat
     7561 tcaacatttc cgtgtcgccc ttattccctt ttttgcggca ttttgccttc ctgtttttgc
     7621 tcacccagaa acgctggtga aagtaaaaga tgctgaagat cagttgggtg cacgagtggg
     7681 ttacatcgaa ctggatctca acagcggtaa gatccttgag agttttcgcc ccgaagaacg
     7741 ttttccaatg atgagcactt ttaaagttct gctatgtggc gcggtattat cccgtattga
     7801 cgccgggcaa gagcaactcg gtcgccgcat acactattct cagaatgact tggttgagta
     7861 ctcaccagtc acagaaaagc atcttacgga tggcatgaca gtaagagaat tatgcagtgc
     7921 tgccataacc atgagtgata acactgcggc caacttactt ctgacaacga tcggaggacc
     7981 gaaggagcta accgcttttt tgcacaacat gggggatcat gtaactcgcc ttgatcgttg
     8041 ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt gacaccacga tgcctgtagc
     8101 aatggcaaca acgttgcgca aactattaac tggcgaacta cttactctag cttcccggca
     8161 acaattaata gactggatgg aggcggataa agttgcagga ccacttctgc gctcggccct
     8221 tccggctggc tggtttattg ctgataaatc tggagccggt gagcgtgggt ctcgcggtat
     8281 cattgcagca ctggggccag atggtaagcc ctcccgtatc gtagttatct acacgacggg
     8341 gagtcaggca actatggatg aacgaaatag acagatcgct gagataggtg cctcactgat
     8401 taagcattgg taactgtcag accaagttta ctcatatata ctttagattg atttaaaact
     8461 tcatttttaa tttaaaagga tctaggtgaa gatccttttt gataatctca tgaccaaaat
     8521 cccttaacgt gagttttcgt tccactgagc gtcagacccc gtagaaaaga tcaaaggatc
     8581 ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct
     8641 accagcggtg gtttgtttgc cggatcaaga gctaccaact ctttttccga aggtaactgg
     8701 cttcagcaga gcgcagatac caaatactgt tcttctagtg tagccgtagt taggccacca
     8761 cttcaagaac tctgtagcac cgcctacata cctcgctctg ctaatcctgt taccagtggc
     8821 tgctgccagt ggcgataagt cgtgtcttac cgggttggac tcaagacgat agttaccgga
     8881 taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca cagcccagct tggagcgaac
     8941 gacctacacc gaactgagat acctacagcg tgagctatga gaaagcgcca cgcttcccga
     9001 agggagaaag gcggacaggt atccggtaag cggcagggtc ggaacaggag agcgcacgag
     9061 ggagcttcca gggggaaacg cctggtatct ttatagtcct gtcgggtttc gccacctctg
     9121 acttgagcgt cgatttttgt gatgctcgtc aggggggcgg agcctatgga aaaacgccag
     9181 caacgcggcc tttttacggt tcctggcctt ttgctggcct tttgctcaca tgttctttcc
     9241 tgcgttatcc cctgattctg tggataaccg tattaccgcc tttgagtgag ctgataccgc
     9301 tcgccgcagc cgaacgaccg agcgcagcga gtcagtgagc gaggaagcgg aagagcgccc
     9361 aatacgcaaa ccgcctctcc ccgcgcgttg gccgattcat taatgcagct ggcacgacag
     9421 gtttcccgac tggaaagcgg gcagtgagcg caacgcaatt aatgtgagtt agctcactca
     9481 ttaggcaccc caggctttac actttatgct tccggctcgt atgttgtgtg gaattgtgag
     9541 cggataacaa tttcacacag gaaacagcta tgaccatgat tacgcc
//
