LOCUS       sequence-356815-        7305 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  TEVP-inducible GFP-conjugated Kir2.1 RELEASE.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7305)
  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 7305)
  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-GFP-Kir2.1-HA-tevs-RXR-V2
FEATURES             Location/Qualifiers
     source          1..7305
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        105..484
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        485..688
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     639..659
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     protein_bind    690..708
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    711..729
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     739..763
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             join(867..869,870..872,873..1583)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     regulatory      1653..1662
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1935..1961
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2922..2942
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             3008..3037
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(3093..3110)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3099..3323
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(3495..3514)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    3646..3693
                     /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             3701..4723
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             3701..4723
                     /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    4853..4974
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4890..4909)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4944..4963
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5023..5039)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5023..5039)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5036..5058)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5047..5063
                     /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(5071..5077,5078..5095,5096..5101))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5116..5137
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5254..5271)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5425..6013)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5505..5524)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6184..6975,6976..7044))
                     /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     6807..6826
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7045..7149)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(7224..7243)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aaaatttaag ctacaacaag gcaaggcttg accgacaatt gcatgaagaa tctgcttagg
       61 gttaggcgtt ttgcgctgct tcgtacgggc cagatatacg cgttgacatt gattattgac
      121 tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata tggagttccg
      181 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
      241 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      301 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      361 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      421 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      481 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      541 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      601 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      661 acggtgggag gtctatataa gcagagctct ccctatcagt gatagagatc tccctatcag
      721 tgatagagat cgtcgacgag ctcgtttagt gaaccgtcag atcgcctgga gacgccatcc
      781 acgctgtttt gacctccata gaagacaccg ggaccgatcc agcctccgga ctctagcgtt
      841 taaacttaag cttcgccacc gctagcatgg tctcaaaggg cgaggaactt tttacgggcg
      901 ttgttccgat cctcgtcgaa ttggacgggg acgttaatgg gcataaattc tcagttagtg
      961 gtgagggtga gggggacgcg acatatggga aacttacgct gaagttcata tgtaccaccg
     1021 gaaaattgcc agtgccgtgg cctactctcg tcaccaccct gacatatggt gttcagtgtt
     1081 ttagccggta tccagaccat atgaaacagc acgatttttt taaatccgct atgcctgaag
     1141 gttacgtaca ggagcgaacg atatttttta aagacgacgg aaactacaaa actagagccg
     1201 aggtaaaatt cgagggtgac actctcgtga accggatcga actgaagggc atagatttta
     1261 aggaggatgg caatatactg ggccacaagc tcgaatacaa ctataattct cataatgtct
     1321 atatcatggc ggacaaacag aaaaacggca tcaaggtaaa ctttaaaatc cgacacaata
     1381 ttgaggacgg ctctgtccag cttgcggacc attaccaaca gaacacgccg ataggggatg
     1441 gtccagtact tctccctgat aatcactatt tgtctactca gtcagctctt agcaaggacc
     1501 caaacgaaaa acgggatcac atggtccttc ttgagttcgt cactgcggct gggataacac
     1561 tcgggatgga cgaactgtat aagggcgcca tgggcagtgt gcgaaccaac cgctacagca
     1621 tcgtctcttc agaagaagac ggtatgaagt tggccaccat ggcagttgca aatggctttg
     1681 ggaacgggaa gagtaaagtc cacacccgac aacagtgcag gagccgcttt gtgaagaaag
     1741 atggccactg taatgttcag ttcatcaatg tgggtgagaa ggggcaacgg tatctcgcag
     1801 acatcttcac cacgtgtgtg gacattcgct ggcggtggat gctggttatc ttctgcctgg
     1861 ctttcgtcct gtcatggctg ttttttggct gtgtgttttg gttgatagct ctgctccatg
     1921 gggacctgga tgcataccca tacgatgttc cagattacgc ttccaaagag ggcaaggctt
     1981 gtgtgtccga ggtcaacagc ttcacggctg ccttcctctt ctccattgag acccagacaa
     2041 ccataggcta tggtttcaga tgtgtcacgg atgaatgccc aattgctgtt ttcatggtgg
     2101 tgttccagtc aatcgtgggc tgcatcatag atgctttcat cattggcgca gtcatggcca
     2161 agatggcaaa gccaaagaag agaaacgaga ctcttgtctt cagtcacaat gccgtgattg
     2221 ccatgagaga cggcaagctg tgtttgatgt ggcgagtggg caatcttcgg aaaagccact
     2281 tggtggaagc tcatgttcga gcacagctcc tcaaatccag aattacttct gaaggggagt
     2341 atatccctct ggatcaaata gacatcaatg ttgggtttga cagtggaata gatcgtatat
     2401 ttctggtgtc cccaatcact atagtccatg aaatagatga agacagtcct ttatatgatt
     2461 tgagtaaaca ggacattgac aacgcagact ttgaaatcgt ggtcatactg gaaggcatgg
     2521 tggaagccac tgccatgacg acacagtgcc gtagctctta tctagcaaat gaaatcctgt
     2581 ggggccaccg ctatgagcct gtgctctttg aagagaagca ctactacaaa gtggactatt
     2641 ccaggttcca caaaacttac gaagtcccca acactcccct ttgtagtgcc agagacttag
     2701 cagaaaagaa atatatcctc tcaaatgcaa attcattttg ctatgaaaat gaagttgccc
     2761 tcacaagcaa agaggaagac gacagtgaaa atggagttcc agaaagcact agtacggaca
     2821 cgccccctga catagacctt cacaaccagg caagtgtacc tctagagccc aggcccttac
     2881 ggcgagagtc ggagatactc gagggcgggg gctcggggct cgaaaacttg tacttccaga
     2941 gtggcagtgg aggtaccaag cacatcctct tccgacggag gagaaggggc ttcaggcagt
     3001 agggcccgaa caaaaactca tctcagaaga ggatctgaat agcgccgtcg accatcatca
     3061 taatcatcat tgagtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc
     3121 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     3181 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct
     3241 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc
     3301 tggggatgcg gtgggctcta tggagatcta tcgatgttct ggggatgcgg tgggctctat
     3361 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
     3421 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     3481 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     3541 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     3601 cctcgacccc aaaaaacttg attagggtga tggttcacgt acctagaagt tcctattccg
     3661 aagttcctat tctctagaaa gtataggaac ttccttggcc aaaaagcctg aactcaccgc
     3721 gacgtctgtc gagaagtttc tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct
     3781 ctcggagggc gaagaatctc gtgctttcag cttcgatgta ggagggcgtg gatatgtcct
     3841 gcgggtaaat agctgcgccg atggtttcta caaagatcgt tatgtttatc ggcactttgc
     3901 atcggccgcg ctcccgattc cggaagtgct tgacattggg gaattcagcg agagcctgac
     3961 ctattgcatc tcccgccgtg cacagggtgt cacgttgcaa gacctgcctg aaaccgaact
     4021 gcccgctgtt ctgcagccgg tcgcggaggc catggatgcg atcgctgcgg ccgatcttag
     4081 ccagacgagc gggttcggcc cattcggacc gcaaggaatc ggtcaataca ctacatggcg
     4141 tgatttcata tgcgcgattg ctgatcccca tgtgtatcac tggcaaactg tgatggacga
     4201 caccgtcagt gcgtccgtcg cgcaggctct cgatgagctg atgctttggg ccgaggactg
     4261 ccccgaagtc cggcacctcg tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa
     4321 tggccgcata acagcggtca ttgactggag cgaggcgatg ttcggggatt cccaatacga
     4381 ggtcgccaac atcttcttct ggaggccgtg gttggcttgt atggagcagc agacgcgcta
     4441 cttcgagcgg aggcatccgg agcttgcagg atcgccgcgg ctccgggcgt atatgctccg
     4501 cattggtctt gaccaactct atcagagctt ggttgacggc aatttcgatg atgcagcttg
     4561 ggcgcagggt cgatgcgacg caatcgtccg atccggagcc gggactgtcg ggcgtacaca
     4621 aatcgcccgc agaagcgcgg ccgtctggac cgatggctgt gtagaagtac tcgccgatag
     4681 tggaaaccga cgccccagca ctcgtccgag ggcaaaggaa tagcacgtac tacgagattt
     4741 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     4801 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt
     4861 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     4921 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     4981 ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
     5041 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     5101 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     5161 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     5221 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     5281 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     5341 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     5401 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     5461 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     5521 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     5581 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     5641 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     5701 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     5761 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     5821 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     5881 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     5941 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     6001 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     6061 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     6121 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     6181 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     6241 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     6301 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     6361 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     6421 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     6481 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     6541 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     6601 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     6661 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     6721 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     6781 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     6841 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     6901 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     6961 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     7021 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     7081 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     7141 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatctccc
     7201 gatcccctat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat
     7261 ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg cgagc
//
