LOCUS       sequence-375413-        6599 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expression of a sensor for synthetic sequence 1 (EK0208) in 
            mammalian cells; mTagBFP2 marker, mCherry output.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6599)
  AUTHORS   Kaseniit KE, Katz N, Kolber NS, Call CC, Wengier DL, Cody WB, 
            Sattely ES, Gao XJ
  TITLE     Modular, programmable RNA sensing using ADAR editing in living 
            cells.
  JOURNAL   Nat Biotechnol. 2022 Oct 5. pii: 10.1038/s41587-022-01493-x. doi: 
            10.1038/s41587-022-01493-x.
  PUBMED    36198772
REFERENCE   2  (bases 1 to 6599)
  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 
            Biotechnol. 2022 Oct 5. pii: 10.1038/s41587-022-01493-x. doi: 
            10.1038/s41587-022-01493-x."
COMMENT     Sequence Label: EK0423 SFFV-mTagBFP2-SGc(s70587)cSG-mCherry (F...
FEATURES             Location/Qualifiers
     source          1..6599
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        112..519
                     /label=SFFV promoter
                     /note="spleen focus-forming virus long terminal repeat 
                     (LTR) promoter"
     regulatory      527..536
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             533..1243
                     /codon_start=1
                     /product="enhanced monomeric blue fluorescent protein 
                     (Subach et al., 2011)"
                     /label=mTagBFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELIKENMHMKLYMEGTVDNHHFKCTSEGEGKPYEGTQTM
                     RIKVVEGGPLPFAFDILATSFLYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGG
                     VLTATQDTSLQDGCLIYNVKIRGVNFTSNGPVMQKKTLGWEAFTETLYPADGGLEGRND
                     MALKLVGGSHLIANAKTTYRSKKPAKNLKMPGVYYVDYRLERIKEANNETYVEQHEVAV
                     ARYCDLPSKLGHKLN"
     CDS             1283..1348
                     /codon_start=1
                     /product="2A peptide from foot-and-mouth disease virus 
                     polyprotein"
                     /label=F2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="VKQTLNFDLLKLAGDVESNPGP"
     CDS             1496..1561
                     /codon_start=1
                     /product="2A peptide from foot-and-mouth disease virus 
                     polyprotein"
                     /label=F2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="VKQTLNFDLLKLAGDVESNPGP"
     CDS             1568..2275
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(1710..1728)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     1978..1997
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2288..2317
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     primer_bind     complement(2373..2390)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2379..2603
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     complement(2775..2794)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     protein_bind    2926..2973
                     /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             2981..4003
                     /codon_start=1
                     /product="hygromycin B phosphotransferase"
                     /label=HygR
                     /note="confers resistance to hygromycin"
                     /translation="KKPELTATSVEKFLIEKFDSVSDLMQLSEGEESRAFSFDVGGRGY
                     VLRVNSCADGFYKDRYVYRHFASAALPIPEVLDIGEFSESLTYCISRRAQGVTLQDLPE
                     TELPAVLQPVAEAMDAIAAADLSQTSGFGPFGPQGIGQYTTWRDFICAIADPHVYHWQT
                     VMDDTVSASVAQALDELMLWAEDCPEVRHLVHADFGSNNVLTDNGRITAVIDWSEAMFG
                     DSQYEVANIFFWRPWLACMEQQTRYFERRHPELAGSPRLRAYMLRIGLDQLYQSLVDGN
                     FDDAAWAQGRCDAIVRSGAGTVGRTQIARRSAAVWTDGCVEVLADSGNRRPSTRPRAKE
                     "
     CDS             2981..4003
                     /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    4133..4254
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4170..4189)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4224..4243
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4303..4319)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4303..4319)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(4316..4338)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    4327..4343
                     /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(4351..4357,4358..4375,4376..4381))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4396..4417
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(4534..4551)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4705..5293)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4785..4804)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5464..6255,6256..6324))
                     /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     6087..6106
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6325..6429)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(6504..6523)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
ORIGIN
        1 aacaaggcaa ggcttgaccg acaattgccg ataaaataaa agattttatt tagtctccag
       61 aaaaaggggg gaatgaaaga ccccacctgt aggtttggca agctagctgc agtaacgcca
      121 ttttgcaagg catggaaaaa taccaaacca agaatagaga agttcagatc aagggcgggt
      181 acatgaaaat agctaacgtt gggccaaaca ggatatctgc ggtgagcagt ttcggccccg
      241 gcccggggcc aagaacagat ggtcaccgca gtttcggccc cggcccgagg ccaagaacag
      301 atggtcccca gatatggccc aaccctcagc agtttcttaa gacccatcag atgtttccag
      361 gctcccccaa ggacctgaaa tgaccctgcg ccttatttga attaaccaat cagcctgctt
      421 ctcgcttctg ttcgcgcgct tctgcttccc gagctctata aaagagctca caacccctca
      481 ctcggcgcgc cagtcctccg acagactgag tcgcccggga agcttcgcca ccatggtgtc
      541 taagggcgaa gagctgatta aggagaacat gcacatgaag ctgtacatgg agggcaccgt
      601 ggacaaccat cacttcaagt gcacatccga gggcgaaggc aagccctacg agggcaccca
      661 gaccatgaga atcaaggtgg tcgagggcgg ccctctcccc ttcgccttcg acatcctggc
      721 tactagcttc ctctacggca gcaagacctt catcaaccac acccagggca tccccgactt
      781 cttcaagcag tccttccctg agggcttcac atgggagaga gtcaccacat acgaggacgg
      841 gggcgtgctg accgctaccc aggacaccag cctccaggac ggctgcctca tctacaacgt
      901 caagatcaga ggggtgaact tcacatccaa cggccctgtg atgcagaaga aaacactcgg
      961 ctgggaggcc ttcaccgaaa cgctgtaccc cgctgacggc ggcctggaag gcagaaacga
     1021 catggccctg aagctcgtgg gcgggagcca tctgatcgca aacgccaaga ccacatatag
     1081 atccaagaaa cccgctaaga acctcaagat gcctggcgtc tactatgtgg actacagact
     1141 ggaaagaatc aaggaggcca acaacgaaac ctacgtcgag cagcacgagg tggcagtggc
     1201 cagatactgc gacctcccta gcaaactggg gcacaaactt aacgcgttgg aagccagaca
     1261 caaacagaaa attgtggctc cggtgaaaca gactttgaat tttgaccttc tgaagctggc
     1321 gggagacgtc gagtccaacc ccggtcccgg taccggatcc accaagtcga cacggatgag
     1381 tgtttccttc tatccgcgtc attgttaggc cttacgtacc gatagaccac tggtgcaaat
     1441 agagagctat accggtacta gtgaggcacg gcataagcaa aagatcgtag ccccagtaaa
     1501 gcaaacactc aacttcgatc tactcaaact cgcaggtgat gtggaatcta atccaggacc
     1561 ctcgagtgtg agcaagggcg aggaggataa catggccatc atcaaggagt tcatgcgctt
     1621 caaggtgcac atggagggct ccgtgaacgg ccacgagttc gagatcgagg gcgagggcga
     1681 gggccgcccc tacgagggca cccagaccgc caagctgaag gtgaccaagg gtggccccct
     1741 gcccttcgcc tgggacatcc tgtcccctca gttcatgtac ggctccaagg cctacgtgaa
     1801 gcaccccgcc gacattcccg actacttgaa gctgtccttc cccgagggct tcaagtggga
     1861 gcgcgtgatg aacttcgagg acggcggcgt ggtgaccgtg acccaggact cctccctgca
     1921 ggacggcgag ttcatctaca aggtgaagct gcgcggcacc aacttcccct ccgacggccc
     1981 cgtaatgcag aagaagacca tgggctggga ggcctcctcc gagcggatgt accccgagga
     2041 cggcgccctg aagggcgaga tcaagcagag gctgaagctg aaggacggcg gccactacga
     2101 cgctgaggtc aagaccacct acaaggccaa gaagcccgtg cagctgcccg gcgcctacaa
     2161 cgtcaacatc aagttggaca tcacctccca caacgaggac tacaccatcg tggaacagta
     2221 cgaacgcgcc gagggccgcc actccaccgg cggcatggac gagctgtaca agtaggccgg
     2281 agggcccgaa caaaaactca tctcagaaga ggatctgaat agcgccgtcg accatcatca
     2341 taatcatcat tgagtttaaa cccgctgatc agcctcgact gtgccttcta gttgccagcc
     2401 atctgttgtt tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt
     2461 cctttcctaa taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct
     2521 ggggggtggg gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc
     2581 tggggatgcg gtgggctcta tggagatcta tcgatgttct ggggatgcgg tgggctctat
     2641 ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg cgccctgtag
     2701 cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag
     2761 cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt
     2821 tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca
     2881 cctcgacccc aaaaaacttg attagggtga tggttcacgt acctagaagt tcctattccg
     2941 aagttcctat tctctagaaa gtataggaac ttccttggcc aaaaagcctg aactcaccgc
     3001 gacgtctgtc gagaagtttc tgatcgaaaa gttcgacagc gtctccgacc tgatgcagct
     3061 ctcggagggc gaagaatctc gtgctttcag cttcgatgta ggagggcgtg gatatgtcct
     3121 gcgggtaaat agctgcgccg atggtttcta caaagatcgt tatgtttatc ggcactttgc
     3181 atcggccgcg ctcccgattc cggaagtgct tgacattggg gaattcagcg agagcctgac
     3241 ctattgcatc tcccgccgtg cacagggtgt cacgttgcaa gacctgcctg aaaccgaact
     3301 gcccgctgtt ctgcagccgg tcgcggaggc catggatgcg atcgctgcgg ccgatcttag
     3361 ccagacgagc gggttcggcc cattcggacc gcaaggaatc ggtcaataca ctacatggcg
     3421 tgatttcata tgcgcgattg ctgatcccca tgtgtatcac tggcaaactg tgatggacga
     3481 caccgtcagt gcgtccgtcg cgcaggctct cgatgagctg atgctttggg ccgaggactg
     3541 ccccgaagtc cggcacctcg tgcacgcgga tttcggctcc aacaatgtcc tgacggacaa
     3601 tggccgcata acagcggtca ttgactggag cgaggcgatg ttcggggatt cccaatacga
     3661 ggtcgccaac atcttcttct ggaggccgtg gttggcttgt atggagcagc agacgcgcta
     3721 cttcgagcgg aggcatccgg agcttgcagg atcgccgcgg ctccgggcgt atatgctccg
     3781 cattggtctt gaccaactct atcagagctt ggttgacggc aatttcgatg atgcagcttg
     3841 ggcgcagggt cgatgcgacg caatcgtccg atccggagcc gggactgtcg ggcgtacaca
     3901 aatcgcccgc agaagcgcgg ccgtctggac cgatggctgt gtagaagtac tcgccgatag
     3961 tggaaaccga cgccccagca ctcgtccgag ggcaaaggaa tagcacgtac tacgagattt
     4021 cgattccacc gccgccttct atgaaaggtt gggcttcgga atcgttttcc gggacgccgg
     4081 ctggatgatc ctccagcgcg gggatctcat gctggagttc ttcgcccacc ccaacttgtt
     4141 tattgcagct tataatggtt acaaataaag caatagcatc acaaatttca caaataaagc
     4201 atttttttca ctgcattcta gttgtggttt gtccaaactc atcaatgtat cttatcatgt
     4261 ctgtataccg tcgacctcta gctagagctt ggcgtaatca tggtcatagc tgtttcctgt
     4321 gtgaaattgt tatccgctca caattccaca caacatacga gccggaagca taaagtgtaa
     4381 agcctggggt gcctaatgag tgagctaact cacattaatt gcgttgcgct cactgcccgc
     4441 tttccagtcg ggaaacctgt cgtgccagct gcattaatga atcggccaac gcgcggggag
     4501 aggcggtttg cgtattgggc gctcttccgc ttcctcgctc actgactcgc tgcgctcggt
     4561 cgttcggctg cggcgagcgg tatcagctca ctcaaaggcg gtaatacggt tatccacaga
     4621 atcaggggat aacgcaggaa agaacatgtg agcaaaaggc cagcaaaagg ccaggaaccg
     4681 taaaaaggcc gcgttgctgg cgtttttcca taggctccgc ccccctgacg agcatcacaa
     4741 aaatcgacgc tcaagtcaga ggtggcgaaa cccgacagga ctataaagat accaggcgtt
     4801 tccccctgga agctccctcg tgcgctctcc tgttccgacc ctgccgctta ccggatacct
     4861 gtccgccttt ctcccttcgg gaagcgtggc gctttctcat agctcacgct gtaggtatct
     4921 cagttcggtg taggtcgttc gctccaagct gggctgtgtg cacgaacccc ccgttcagcc
     4981 cgaccgctgc gccttatccg gtaactatcg tcttgagtcc aacccggtaa gacacgactt
     5041 atcgccactg gcagcagcca ctggtaacag gattagcaga gcgaggtatg taggcggtgc
     5101 tacagagttc ttgaagtggt ggcctaacta cggctacact agaagaacag tatttggtat
     5161 ctgcgctctg ctgaagccag ttaccttcgg aaaaagagtt ggtagctctt gatccggcaa
     5221 acaaaccacc gctggtagcg gtggtttttt tgtttgcaag cagcagatta cgcgcagaaa
     5281 aaaaggatct caagaagatc ctttgatctt ttctacgggg tctgacgctc agtggaacga
     5341 aaactcacgt taagggattt tggtcatgag attatcaaaa aggatcttca cctagatcct
     5401 tttaaattaa aaatgaagtt ttaaatcaat ctaaagtata tatgagtaaa cttggtctga
     5461 cagttaccaa tgcttaatca gtgaggcacc tatctcagcg atctgtctat ttcgttcatc
     5521 catagttgcc tgactccccg tcgtgtagat aactacgata cgggagggct taccatctgg
     5581 ccccagtgct gcaatgatac cgcgagaccc acgctcaccg gctccagatt tatcagcaat
     5641 aaaccagcca gccggaaggg ccgagcgcag aagtggtcct gcaactttat ccgcctccat
     5701 ccagtctatt aattgttgcc gggaagctag agtaagtagt tcgccagtta atagtttgcg
     5761 caacgttgtt gccattgcta caggcatcgt ggtgtcacgc tcgtcgtttg gtatggcttc
     5821 attcagctcc ggttcccaac gatcaaggcg agttacatga tcccccatgt tgtgcaaaaa
     5881 agcggttagc tccttcggtc ctccgatcgt tgtcagaagt aagttggccg cagtgttatc
     5941 actcatggtt atggcagcac tgcataattc tcttactgtc atgccatccg taagatgctt
     6001 ttctgtgact ggtgagtact caaccaagtc attctgagaa tagtgtatgc ggcgaccgag
     6061 ttgctcttgc ccggcgtcaa tacgggataa taccgcgcca catagcagaa ctttaaaagt
     6121 gctcatcatt ggaaaacgtt cttcggggcg aaaactctca aggatcttac cgctgttgag
     6181 atccagttcg atgtaaccca ctcgtgcacc caactgatct tcagcatctt ttactttcac
     6241 cagcgtttct gggtgagcaa aaacaggaag gcaaaatgcc gcaaaaaagg gaataagggc
     6301 gacacggaaa tgttgaatac tcatactctt cctttttcaa tattattgaa gcatttatca
     6361 gggttattgt ctcatgagcg gatacatatt tgaatgtatt tagaaaaata aacaaatagg
     6421 ggttccgcgc acatttcccc gaaaagtgcc acctgacgtc gacggatcgg gagatctccc
     6481 gatcccctat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagtat
     6541 ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg cgagcaaaat ttaagctac
//
