LOCUS       sequence-371944-        4021 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Fluorescent reporter for cAMP.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4021)
  AUTHORS   Kawata S, Mukai Y, Nishimura Y, Takahashi T, Saitoh N
  TITLE     Green fluorescent cAMP indicator of high speed and specificity 
            suitable for neuronal live-cell imaging.
  JOURNAL   Proc Natl Acad Sci U S A. 2022 Jul 12;119(28):e2122618119. doi: 
            10.1073/pnas.2122618119. Epub 2022 Jul 6.
  PUBMED    35867738
REFERENCE   2  (bases 1 to 4021)
  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: "Proc Natl 
            Acad Sci U S A."; date: "2022-07-12"; volume: "119(28):e2122618119. 
            doi"; pages: " 10.1073/pnas.2122618119. Epub 2022 Jul 6"
COMMENT     Sequence Label: pUC57-gCarvi-T2A
FEATURES             Location/Qualifiers
     source          1..4021
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(29..51)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     151..170
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     364..386
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     primer_bind     378..395
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     379..395
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     CDS             complement(471..524)
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     primer_bind     658..677
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     complement(1044..1065)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             complement(1692..1757)
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     regulatory      1757..1766
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     primer_bind     complement(1800..1816)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1800..1816)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1813..1835)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1824..1840
                     /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(1848..1854,1855..1872,1873..1878))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1893..1914
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(2031..2048)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2202..2790)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2282..2301)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2961..3752,3753..3821))
                     /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     3584..3603
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3822..3926)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     3994..4012
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
ORIGIN
        1 tcgcgcgttt cggtgatgac ggtgaaaacc tctgacacat gcagctcccg gagacggtca
       61 cagcttgtct gtaagcggat gccgggagca gacaagcccg tcagggcgcg tcagcgggtg
      121 ttggcgggtg tcggggctgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
      181 accatatgcg gtgtgaaata ccgcacagat gcgtaaggag aaaataccgc atcaggcgcc
      241 attcgccatt caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat
      301 tacgccagct ggcgaaaggg ggatgtgctg caaggcgatt aagttgggta acgccagggt
      361 tttcccagtc acgacgttgt aaaacgacgg ccagtgaatt cgagctcggt acctcgcgaa
      421 tgcatctaga tcccaatgtc gacggtatcg ataagcttga tatcgaattc agggccaggg
      481 ttttcctcca cgtcgccgca ggtcagcagg ctgcctctgc cctcagatcc ctcgacaagc
      541 ttactagtgt tgtactccag cttgtgcccc aggatgttgc cgtcctcctt gaagtcgatg
      601 cccttcagct cgatgcggtt caccagggtg tcgccctcga acttcacctc ggcgcgggtc
      661 ttgtagttgc cgtcgtcctt gaagaagatg gtgcgctcct ggatgtagcc ttcgggcatg
      721 gcggacttga agaagtcgtg ctgcttcatg tggtcggggt agcggctgaa gcactgcacg
      781 ccgtaggtca gggtggtcac gagggtgggc cagggcacgg gcagcttgcc ggtggtgcag
      841 atgaacttca gggtcagctt gccgtaggtg gcatcgccct caccctcgcc ggacacgctg
      901 aacttgtggc cgtttacgtc gccgtccagc tcgacctgga tgggcaccac cccggtgaac
      961 agctcctcgc ccttgctcac catgctccct ccggtaccgc ccttgtacag ctcgtccatg
     1021 ccgagagtga tcccggcggc ggtcacgaac tccagcagga ccatgtgatc gcgcttctcg
     1081 ttggggtctt tcgaaagaat ggactgcacg ctcaggtagt ggttgtcggg cagcagcacg
     1141 gggccgtcgc cgatgggggt gttctgctgg tagtggtagg cgagctgcac gccgccgtcc
     1201 tcgatgttgt ggcggatctt gaagttcgcc ttgatgccgt tcttctgctt gtcggccttg
     1261 atatagacgt tctcgaggta caggttcccc accttctcag aggtaacctg aagccttctc
     1321 gccatctggg ctgagaggcg catgagtatg tcgggattca cctgtatgag ttgccggaat
     1381 ttcttgtatg agatttcggc cacctcacag gctgtcttag ccctcaccca agcggatctc
     1441 tcctgtcctt cctcgaacag gcccagttct ccgataaagt caccctggtt cagatagctc
     1501 aggatcattt cctttccttc ttcatcctta atcagcacgg cgactgaacc cttcactata
     1561 taatacaggg tttcagcctt ctcgccctga tggatcaggg tgctcttaga tgggtacttg
     1621 tgaatgtgac agtgtgacag aaaccactcc aatgttgggt cggtctgtgg tttgcccagc
     1681 accatggatc ccttgtcatc gtcatccttg taatcgatgt catgatcttt ataatcaccg
     1741 tcatggtctt tgtagtccat ggtggcacta gtagatcact agttctagta gatctcatgg
     1801 tcatagctgt ttcctgtgtg aaattgttat ccgctcacaa ttccacacaa catacgagcc
     1861 ggaagcataa agtgtaaagc ctggggtgcc taatgagtga gctaactcac attaattgcg
     1921 ttgcgctcac tgcccgcttt ccagtcggga aacctgtcgt gccagctgca ttaatgaatc
     1981 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     2041 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     2101 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     2161 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     2221 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     2281 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     2341 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     2401 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     2461 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     2521 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     2581 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     2641 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     2701 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     2761 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     2821 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     2881 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     2941 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     3001 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     3061 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     3121 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     3181 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     3241 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     3301 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     3361 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     3421 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     3481 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     3541 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     3601 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     3661 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     3721 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     3781 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     3841 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     3901 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc tgacgtctaa
     3961 gaaaccatta ttatcatgac attaacctat aaaaataggc gtatcacgag gccctttcgt
     4021 c
//
