LOCUS       sequence-355662-        7032 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  OR Gate (TEVP/HCVP) SEAP RELEASE construct.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7032)
  AUTHORS   Vlahos AE, Kang J, Aldrete CA, Zhu R, Chong LS, Elowitz MB, Gao XJ
  TITLE     Protease-controlled secretion and display of intercellular signals.
  JOURNAL   Nat Commun. 2022 Feb 17;13(1):912. doi: 10.1038/s41467-022-28623-y.
  PUBMED    35177637
REFERENCE   2  (bases 1 to 7032)
  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: "Nat 
            Commun."; date: "2022-02-17"; volume: "13(1):912. doi"; pages: " 
            10.1038/s41467-022-28623-y"
COMMENT     Sequence Label: CMVTO-SEAP-26sFur-B2AD-hcvs-tevs-KKMP
FEATURES             Location/Qualifiers
     source          1..7032
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     620..639
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(858..962)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(1037..1056)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        1223..1602
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        1603..1806
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1757..1777
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    1808..1826
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1829..1847
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1857..1881
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             1979..3481
                     /codon_start=1
                     /product="secreted alkaline phosphatase from human 
                     placenta"
                     /label=SEAP
                     /note="The natural enzyme has a C-terminal transmembrane 
                     sequence that is replaced by a GPI anchor."
                     /translation="MLLLLLLLGLRLQLSLGIIPVEEENPDFWNREAAEALGAAKKLQP
                     AQTAAKNLIIFLGDGMGVSTVTAARILKGQKKDKLGPEIPLAMDRFPYVALSKTYNVDK
                     HVPDSGATATAYLCGVKGNFQTIGLSAAARFNQCNTTRGNEVISVMNRAKKAGKSVGVV
                     TTTRVQHASPAGTYAHTVNRNWYSDADVPASARQEGCQDIATQLISNMDIDVILGGGRK
                     YMFPMGTPDPEYPDDYSQGGTRLDGKNLVQEWLAKRQGARYVWNRTELMQASLDPSVTH
                     LMGLFEPGDMKYEIHRDSTLDPSLMEMTEAALRLLSRNPRGFFLFVEGGRIDHGHHESR
                     AYRALTETIMFDDAIERAGQLTSEEDTLSLVTADHSHVFSFGGYPLRGSSIFGLAPGKA
                     RDRKAYTVLLYGNGPGYVLKDGARPDVTESESGSPEYRQQSAVPLDEETHAGEDVAVFA
                     RGPQAHLVHGVQEQTFIAHVMAFAACLEPYTACDLAPPAGTTD"
     CDS             3764..3784
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             3853..3882
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(3938..3955)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3944..4168
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(4340..4359)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    4491..4538
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             4546..5568
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             4546..5568
                     /codon_start=1
                     /gene="aph(4)-Ia"
                     /product="aminoglycoside phosphotransferase from E. coli"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     polyA_signal    5698..5819
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5735..5754)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5789..5808
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5868..5884)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5868..5884)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5881..5903)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5892..5908
                     /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(5916..5922,5923..5940,5941..5946))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5961..5982
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(6099..6116)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(6270..6858)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(6350..6369)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7029..7032,1..788,789..857))
                     /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"
ORIGIN
        1 caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc atccatagtt
       61 gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc tggccccagt
      121 gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc aataaaccag
      181 ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc catccagtct
      241 attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt gcgcaacgtt
      301 gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc ttcattcagc
      361 tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa aaaagcggtt
      421 agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt atcactcatg
      481 gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg cttttctgtg
      541 actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc gagttgctct
      601 tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa agtgctcatc
      661 attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt gagatccagt
      721 tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt caccagcgtt
      781 tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag ggcgacacgg
      841 aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta tcagggttat
      901 tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat aggggttccg
      961 cgcacatttc cccgaaaagt gccacctgac gtcgacggat cgggagatct cccgatcccc
     1021 tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag tatctgctcc
     1081 ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa aatttaagct acaacaaggc
     1141 aaggcttgac cgacaattgc atgaagaatc tgcttagggt taggcgtttt gcgctgcttc
     1201 gtacgggcca gatatacgcg ttgacattga ttattgacta gttattaata gtaatcaatt
     1261 acggggtcat tagttcatag cccatatatg gagttccgcg ttacataact tacggtaaat
     1321 ggcccgcctg gctgaccgcc caacgacccc cgcccattga cgtcaataat gacgtatgtt
     1381 cccatagtaa cgccaatagg gactttccat tgacgtcaat gggtggagta tttacggtaa
     1441 actgcccact tggcagtaca tcaagtgtat catatgccaa gtacgccccc tattgacgtc
     1501 aatgacggta aatggcccgc ctggcattat gcccagtaca tgaccttatg ggactttcct
     1561 acttggcagt acatctacgt attagtcatc gctattacca tggtgatgcg gttttggcag
     1621 tacatcaatg ggcgtggata gcggtttgac tcacggggat ttccaagtct ccaccccatt
     1681 gacgtcaatg ggagtttgtt ttggcaccaa aatcaacggg actttccaaa atgtcgtaac
     1741 aactccgccc cattgacgca aatgggcggt aggcgtgtac ggtgggaggt ctatataagc
     1801 agagctctcc ctatcagtga tagagatctc cctatcagtg atagagatcg tcgacgagct
     1861 cgtttagtga accgtcagat cgcctggaga cgccatccac gctgttttga cctccataga
     1921 agacaccggg accgatccag cctccggact ctagcgttta aacttaagct tcgccaccat
     1981 gctgctcctg ttgctgctgc tgggcctgag gctacagctc tccctgggca tcatcccagt
     2041 tgaggaggag aacccggact tctggaaccg cgaggcagcc gaggccctgg gtgccgccaa
     2101 gaagctgcag cctgcacaga cagccgccaa gaacctcatc atcttcctgg gcgatgggat
     2161 gggggtgtct acggtgacag ctgccaggat tctaaaaggg cagaagaagg acaaactggg
     2221 gcctgagata cccctggcca tggaccgctt cccatatgtg gctctgtcca agacatacaa
     2281 tgtagacaaa catgtgccag acagtggagc cacagccacg gcctacctgt gcggggtcaa
     2341 gggcaacttc cagaccattg gcttgagtgc agccgcccgc tttaaccagt gcaacacgac
     2401 acgcggcaac gaggtcatct ccgtgatgaa tcgggccaag aaagcaggga agtcagtggg
     2461 agtggtaacc accacacgag tgcagcacgc ctcgccagcc ggcacctacg cccacacggt
     2521 gaaccgcaac tggtactcgg acgccgacgt gcctgcctcg gcccgccagg aggggtgcca
     2581 ggacatcgct acgcagctca tctccaacat ggacattgac gtgatcctag gtggaggccg
     2641 aaaatacatg tttcccatgg gaaccccaga ccctgagtac ccagatgact acagccaagg
     2701 tgggaccagg ctggacggga agaatctggt gcaggaatgg ctggcgaagc gccagggtgc
     2761 ccggtatgtg tggaaccgca ctgagctcat gcaggcttcc ctggacccgt ctgtgaccca
     2821 tctcatgggt ctctttgagc ctggagacat gaaatacgag atccaccgag actccacact
     2881 ggacccctcc ctgatggaga tgacagaggc tgccctgcgc ctgctgagca ggaacccccg
     2941 cggcttcttc ctcttcgtgg agggtggtcg catcgaccat ggtcatcatg aaagcagggc
     3001 ttaccgggca ctgactgaga cgatcatgtt cgacgacgcc attgagaggg cgggccagct
     3061 caccagcgag gaggacacgc tgagcctcgt cactgccgac cactcccacg tcttctcctt
     3121 cggaggctac cccctgcgag ggagctccat cttcgggctg gcccctggca aggcccggga
     3181 caggaaggcc tacacggtcc tcctatacgg aaacggtcca ggctatgtgc tcaaggacgg
     3241 cgctcggccg gatgttaccg agagcgagag cgggagcccc gagtatcggc agcagtcagc
     3301 agtgcccctg gacgaagaga cccacgcagg cgaggacgtg gcggtgttcg cgcgcggccc
     3361 gcaggcgcac ctggttcacg gcgtgcagga gcagaccttc atagcgcacg tcatggcctt
     3421 cgccgcctgc ctggagccct acaccgcctg cgacctggca ccccccgccg gcaccaccga
     3481 cgccgcgcac ccgggtggat cctctggcag aagcaaaaga agcgtcattg acggctttac
     3541 cctgaccagc gacgaggtgt gggtggtggg catgggcatc gtcatgtctc tcatcgtcct
     3601 ggccatcgtg tttggcaatg tgctggtcat cacagccatt gccaagttcg agcgtctgca
     3661 gacggtcacc tctaccgtga gcgaggaagc tagtgaggat gtcgtctgct gctcaatgtc
     3721 ctacacatgg acaggagccc tgggtaccgg gggcggctct ctcgagaact tgtacttcca
     3781 gagtggcagt ggaggtacca agtacaaaag cagacgcagt tttattgatg aaaagaaaat
     3841 gccttagggc ccgaacaaaa actcatctca gaagaggatc tgaatagcgc cgtcgaccat
     3901 catcataatc atcattgagt ttaaacccgc tgatcagcct cgactgtgcc ttctagttgc
     3961 cagccatctg ttgtttgccc ctcccccgtg ccttccttga ccctggaagg tgccactccc
     4021 actgtccttt cctaataaaa tgaggaaatt gcatcgcatt gtctgagtag gtgtcattct
     4081 attctggggg gtggggtggg gcaggacagc aagggggagg attgggaaga caatagcagg
     4141 catgctgggg atgcggtggg ctctatggag atctatcgat gttctgggga tgcggtgggc
     4201 tctatggctt ctgaggcgga aagaaccagc tggggctcta gggggtatcc ccacgcgccc
     4261 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     4321 gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     4381 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     4441 cggcacctcg accccaaaaa acttgattag ggtgatggtt cacgtaccta gaagttccta
     4501 ttccgaagtt cctattctct agaaagtata ggaacttcct tggccaaaaa gcctgaactc
     4561 accgcgacgt ctgtcgagaa gtttctgatc gaaaagttcg acagcgtctc cgacctgatg
     4621 cagctctcgg agggcgaaga atctcgtgct ttcagcttcg atgtaggagg gcgtggatat
     4681 gtcctgcggg taaatagctg cgccgatggt ttctacaaag atcgttatgt ttatcggcac
     4741 tttgcatcgg ccgcgctccc gattccggaa gtgcttgaca ttggggaatt cagcgagagc
     4801 ctgacctatt gcatctcccg ccgtgcacag ggtgtcacgt tgcaagacct gcctgaaacc
     4861 gaactgcccg ctgttctgca gccggtcgcg gaggccatgg atgcgatcgc tgcggccgat
     4921 cttagccaga cgagcgggtt cggcccattc ggaccgcaag gaatcggtca atacactaca
     4981 tggcgtgatt tcatatgcgc gattgctgat ccccatgtgt atcactggca aactgtgatg
     5041 gacgacaccg tcagtgcgtc cgtcgcgcag gctctcgatg agctgatgct ttgggccgag
     5101 gactgccccg aagtccggca cctcgtgcac gcggatttcg gctccaacaa tgtcctgacg
     5161 gacaatggcc gcataacagc ggtcattgac tggagcgagg cgatgttcgg ggattcccaa
     5221 tacgaggtcg ccaacatctt cttctggagg ccgtggttgg cttgtatgga gcagcagacg
     5281 cgctacttcg agcggaggca tccggagctt gcaggatcgc cgcggctccg ggcgtatatg
     5341 ctccgcattg gtcttgacca actctatcag agcttggttg acggcaattt cgatgatgca
     5401 gcttgggcgc agggtcgatg cgacgcaatc gtccgatccg gagccgggac tgtcgggcgt
     5461 acacaaatcg cccgcagaag cgcggccgtc tggaccgatg gctgtgtaga agtactcgcc
     5521 gatagtggaa accgacgccc cagcactcgt ccgagggcaa aggaatagca cgtactacga
     5581 gatttcgatt ccaccgccgc cttctatgaa aggttgggct tcggaatcgt tttccgggac
     5641 gccggctgga tgatcctcca gcgcggggat ctcatgctgg agttcttcgc ccaccccaac
     5701 ttgtttattg cagcttataa tggttacaaa taaagcaata gcatcacaaa tttcacaaat
     5761 aaagcatttt tttcactgca ttctagttgt ggtttgtcca aactcatcaa tgtatcttat
     5821 catgtctgta taccgtcgac ctctagctag agcttggcgt aatcatggtc atagctgttt
     5881 cctgtgtgaa attgttatcc gctcacaatt ccacacaaca tacgagccgg aagcataaag
     5941 tgtaaagcct ggggtgccta atgagtgagc taactcacat taattgcgtt gcgctcactg
     6001 cccgctttcc agtcgggaaa cctgtcgtgc cagctgcatt aatgaatcgg ccaacgcgcg
     6061 gggagaggcg gtttgcgtat tgggcgctct tccgcttcct cgctcactga ctcgctgcgc
     6121 tcggtcgttc ggctgcggcg agcggtatca gctcactcaa aggcggtaat acggttatcc
     6181 acagaatcag gggataacgc aggaaagaac atgtgagcaa aaggccagca aaaggccagg
     6241 aaccgtaaaa aggccgcgtt gctggcgttt ttccataggc tccgcccccc tgacgagcat
     6301 cacaaaaatc gacgctcaag tcagaggtgg cgaaacccga caggactata aagataccag
     6361 gcgtttcccc ctggaagctc cctcgtgcgc tctcctgttc cgaccctgcc gcttaccgga
     6421 tacctgtccg cctttctccc ttcgggaagc gtggcgcttt ctcatagctc acgctgtagg
     6481 tatctcagtt cggtgtaggt cgttcgctcc aagctgggct gtgtgcacga accccccgtt
     6541 cagcccgacc gctgcgcctt atccggtaac tatcgtcttg agtccaaccc ggtaagacac
     6601 gacttatcgc cactggcagc agccactggt aacaggatta gcagagcgag gtatgtaggc
     6661 ggtgctacag agttcttgaa gtggtggcct aactacggct acactagaag aacagtattt
     6721 ggtatctgcg ctctgctgaa gccagttacc ttcggaaaaa gagttggtag ctcttgatcc
     6781 ggcaaacaaa ccaccgctgg tagcggtggt ttttttgttt gcaagcagca gattacgcgc
     6841 agaaaaaaag gatctcaaga agatcctttg atcttttcta cggggtctga cgctcagtgg
     6901 aacgaaaact cacgttaagg gattttggtc atgagattat caaaaaggat cttcacctag
     6961 atccttttaa attaaaaatg aagttttaaa tcaatctaaa gtatatatga gtaaacttgg
     7021 tctgacagtt ac
//
