LOCUS       sequence-230378-        8991 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  ddRFP-based single-color biosensor for monitoring cAMP dynamics..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8991)
  AUTHORS   Mehta S, Zhang Y, Roth RH, Zhang JF, Mo A, Tenner B, Huganir RL, 
            Zhang J
  TITLE     Single-fluorophore biosensors for sensitive and multiplexed 
            detection of signalling activities.
  JOURNAL   Nat Cell Biol. 2018 Sep 24. pii: 10.1038/s41556-018-0200-6. doi: 
            10.1038/s41556-018-0200-6.
  PUBMED    30250062
REFERENCE   2  (bases 1 to 8991)
  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 Cell 
            Biol. 2018 Sep 24. pii: 10.1038/s41556-018-0200-6. doi: 
            10.1038/s41556-018-0200-6."
COMMENT     Sequence Label: pcDNA3-RAB-ICUE
FEATURES             Location/Qualifiers
     source          1..8991
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(44..63)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        235..614
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        615..818
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     769..789
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     863..882
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        863..881
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      936..945
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(4582..4599)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4588..4812
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      4858..5286
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4945..4964)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5155..5176
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5295..5315)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        5300..5629
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      5480..5615
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     5542..5561
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             5696..6490
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(5750..5769)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     6360..6379
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    6664..6785
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6701..6720)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6755..6774
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(6834..6850)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(6834..6850)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(6847..6869)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    6858..6874
                     /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(6882..6888,6889..6906,6907..6912))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    6927..6948
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(7065..7082)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(7236..7824)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(7316..7335)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(7995..8786,8787..8855))
                     /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     8618..8637
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(8856..8960)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gacggatcgg gagatctccc gatcccctat ggtgcactct cagtacaatc tgctctgatg
       61 ccgcatagtt aagccagtat ctgctccctg cttgtgtgtt ggaggtcgct gagtagtgcg
      121 cgagcaaaat ttaagctaca acaaggcaag gcttgaccga caattgcatg aagaatctgc
      181 ttagggttag gcgttttgcg ctgcttcgcg atgtacgggc cagatatacg cgttgacatt
      241 gattattgac tagttattaa tagtaatcaa ttacggggtc attagttcat agcccatata
      301 tggagttccg cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc
      361 cccgcccatt gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc
      421 attgacgtca atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt
      481 atcatatgcc aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt
      541 atgcccagta catgacctta tgggactttc ctacttggca gtacatctac gtattagtca
      601 tcgctattac catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg
      661 actcacgggg atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc
      721 aaaatcaacg ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg
      781 gtaggcgtgt acggtgggag gtctatataa gcagagctct ctggctaact agagaaccca
      841 ctgcttactg gcttatcgaa attaatacga ctcactatag ggagacccaa gctggctagc
      901 gtttaaactt aagcttggta ccgagctcgg atccggccac catggtgagc aagagcgagg
      961 aggtcatcaa agagttcatg cgcttcaagg tgcgcttgga gggctccatg aacggccacg
     1021 agttcgagat cgagggcgag ggcgagggcc gcccctacga gggcacccag accgccaagc
     1081 tgaaggtgac caagggcggc cccctgccct tcgcctggga catcctgtcc ccccagatca
     1141 tgtacggctc caaggcgtac gtgaagcacc ccgccgacgt ccccgattac atgaagctgt
     1201 ccttccccga gggcttcaag tgggagcgcg tgatgcactt cgaggacggc ggtctggtga
     1261 ccgtgactca ggacacctcc ctgcaggacg gcacgctgat ctacaaggtg aagatgcgcg
     1321 gcaccaactt cccccccgac ggccccgtaa tgcagaagaa gaccttgggc tgggattatg
     1381 ccaccgagcg cctgtacccc gaagacggcg tgctgaaggg cgagcttctg gggcgcctga
     1441 agctgaagga cggcggcctc aacctggtgg agttcaagac catctacatg gccaagaagc
     1501 ccgtgcaact gcccggctac tacttcgtgg acaccaagct gggcatcacc tcccacaacg
     1561 aggactacac catcgtggaa cagtacgagc gctccgaggg ccgccaccac ctggggcatg
     1621 gcaccggcag cgcatgcccc gtgggaactc atgagatgga ggaggagttg gccgaagctg
     1681 tggccctgct ctcccagcgg gggcctgacg ccctgctcac tgtggcactt cgaaagcccc
     1741 caggtcagcg cacggatgaa gagctggacc tcatctttga ggagctgctg cacatcaagg
     1801 ctgtggccca cctctccaac tcggtgaagc gagaattagc ggctgttctg ctctttgaac
     1861 cacacagcaa ggcagggacc gtgttgttca gccaggggga caagggcact tcgtggtaca
     1921 ttatctggaa gggatctgtc aacgtggtga cccatggcaa ggggctggtg accaccctgc
     1981 atgagggaga tgattttgga cagctggctc tggtgaatga tgcaccccgg gcagccacca
     2041 tcatcctgcg agaagacaac tgtcatttcc tgcgtgtgga caagcaggac ttcaaccgta
     2101 tcatcaagga tgtggaggca aagaccatgc ggctggaaga acatggcaaa gtggtgctgg
     2161 tgctggagag agcctctcag ggcgccggcc cttcccgacc cccaacccca ggcaggaacc
     2221 ggtatacagt gatgtctggc accccagaga agatcctaga gcttctgttg gaggccatgg
     2281 gaccagattc cagtgctcat gaccaaacag agacattcct cagcgacttc ctcctgaccc
     2341 acagggtctt catgcccagc gcccaactct gcgctgccct tctgcaccac ttccatgtgg
     2401 agcctgcggg tggcagcgag caggagcgca gcacctacgt ctgcaacaag aggcagcaga
     2461 tcttgcggct ggtcagccag tgggtggccc tgtatggctc catgctccac actgaccctg
     2521 tggccaccag cttcctccag aaactctcag acctggtggg cagggacacc cgactcagca
     2581 acctgctgag ggagcagtgg ccagagaggc ggcgatgcca caggttggag aatggctgtg
     2641 ggaatgcatc tcctcagatg aaggcccgga acttgcctgt ttggctcccc aaccaggacg
     2701 agccccttcc tggcagcagc tgtgccatcc aagttgggga taaagtcccc tatgacatct
     2761 gccggccaga ccactcagtg ttgaccctgc agctgcctgt gacagcctcc gtgagagagg
     2821 tgatggcagc gttggcccag gaggatggct ggaccaaggg gcaggtgctg gtgaaggtca
     2881 attctgcagg tgatgccatt ggcctgcagc cagatgcccg tggtgtggcc acatctctgg
     2941 ggctcaatga gcgtctcttt gttgtcaacc cacaggaagc gcatgagctg atcccacacc
     3001 ctgaccagct ggggcccact gtgggctctg ctgaggggct ggacctggtg agtgccaagg
     3061 acctggcagg ccagctgacg gaccacgact ggagcctctt caacagtatc caccaggtgg
     3121 agctgatcca ctatgtgctg ggcccccagc atctgcggga tgtcaccacc gccaacctgg
     3181 agcgcttcat gcgccgcttc aatgagctgc agtactgggt ggccaccgag ctgtgtctct
     3241 gccccgtgcc cggcccccgg gcccagctgc tcaggaagtt cattaagctg gcggcccacc
     3301 tcaaggagca gaagaatctc aattccttct ttgccgtcat gtttggcctc agcaactcgg
     3361 ccatcagcca cctagcccac acctgggagc ggctgcccca caaagtccgg aagctgtact
     3421 ccgccctcga gaggctgctg gacccctcat ggaaccaccg ggtataccga ctggccctcg
     3481 ccaagctctc ccctcctgtc atccccttca tgccccttct tctcaaagac atgaccttca
     3541 ttcatgaggg aaaccacaca ctagtggaga atctcatcaa ctttgagaag atgagaatga
     3601 tggccagagt cgcgcggatg ctgcaccact gccgaagcca caaccctgtg cctctctcac
     3661 cactcagaag ccgagtttcc cacctccacg aggacagcca ggtggcgagg atttccacat
     3721 gctcggagca gtccctgagc acccggagtc cagccagcac ctgggcttat gtccagcagc
     3781 tgaaggtcat tgacaaccag cgggaactct cccgcctctc ccgagagctg gagccagagc
     3841 tcgcctcctc cgaggacaac aacatggcca ccatcaaaga gttcatgcgc ttcaaggtgc
     3901 gcatggaggg ctccatgaac ggccacgagt tcgagatcga gggcgagggc gagggccgcc
     3961 cctacgaggg cacccagacc gccaagctga aggtgaccaa gggcggcccc ctgcccttcg
     4021 cctgggacat cctgtccccc cagttcatgt acggctccga ggcgtacgtg aggcaccccg
     4081 ccgacatccc cgattacaag aagctgccct tccccgaggg cttcaagtgg gagcgcgtga
     4141 tgaacttcga agacggcggt ctggtgaccg ttacccagga ctcctccctg caggacggca
     4201 cgctgatctg caaggtgaag atgcgcggca ccaacttccc ccccgacggc cccgtaatgc
     4261 agaagaagac catgggctgg gaggcctcca ccgagatgct gtaccccgaa gacggcgtgc
     4321 tgaagggcca tagctatcag gccctgaagc tgaaggacgg cggccactac ctggtggagt
     4381 tcgagaccat ctacatggcc aagaagcccg tgcaactgcc cggcgattac tgtgtggaca
     4441 ccaagctgga catcacctcc cacaacgagg actacaccat cgtggaacag tacgggcgct
     4501 ccgagggccg ccaccgtctg ggcatggacg agcggtacaa ggaattctaa tctagagggc
     4561 ccgtttaaac ccgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     4621 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     4681 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     4741 tggggcagga cagcaagggg gaggattggg aagacaatag caggcatgct ggggatgcgg
     4801 tgggctctat ggcttctgag gcggaaagaa ccagctgggg ctctaggggg tatccccacg
     4861 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
     4921 cacttgccag cgccctagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
     4981 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
     5041 ctttacggca cctcgacccc aaaaaacttg attagggtga tggttcacgt agtgggccat
     5101 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
     5161 tcttgttcca aactggaaca acactcaacc ctatctcggt ctattctttt gatttataag
     5221 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     5281 cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
     5341 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
     5401 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
     5461 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
     5521 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
     5581 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
     5641 agcttgtata tccattttcg gatctgatca agagacagga tgaggatcgt ttcgcatgat
     5701 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
     5761 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
     5821 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga
     5881 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
     5941 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
     6001 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
     6061 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
     6121 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
     6181 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga
     6241 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
     6301 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
     6361 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
     6421 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
     6481 gttcttctga gcgggactct ggggttcgaa atgaccgacc aagcgacgcc caacctgcca
     6541 tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg aatcgttttc
     6601 cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt cttcgcccac
     6661 cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat cacaaatttc
     6721 acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact catcaatgta
     6781 tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc atggtcatag
     6841 ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg agccggaagc
     6901 ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat tgcgttgcgc
     6961 tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg aatcggccaa
     7021 cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg
     7081 ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg
     7141 ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag
     7201 gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac
     7261 gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga
     7321 taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt
     7381 accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc
     7441 tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc
     7501 cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta
     7561 agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat
     7621 gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca
     7681 gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct
     7741 tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt
     7801 acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct
     7861 cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc
     7921 acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa
     7981 acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta
     8041 tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc
     8101 ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat
     8161 ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta
     8221 tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt
     8281 aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt
     8341 ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg
     8401 ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc
     8461 gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc
     8521 gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg
     8581 cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga
     8641 actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta
     8701 ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct
     8761 tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag
     8821 ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca atattattga
     8881 agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat
     8941 aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt c
//
