LOCUS       sequence-313715-        6458 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  2P-activatable bPAC for transient expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6458)
  AUTHORS   Kinjo T, Watabe T, Kobachi K, Terai K, Matsuda M
  TITLE     Single-Cell Activation of the cAMP-Signaling Pathway in 3D Tissues 
            with FRET-Assisted Two-Photon Activation of bPAC.
  JOURNAL   ACS Chem Biol. 2020 Oct 19. doi: 10.1021/acschembio.0c00333.
  PUBMED    33074647
REFERENCE   2  (bases 1 to 6458)
  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: "ACS Chem 
            Biol. 2020 Oct 19. doi: 10.1021/acschembio.0c00333."
COMMENT     Sequence Label: pCAGGS_2pabPAC
FEATURES             Location/Qualifiers
     source          1..6458
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        9..388
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     intron          666..1686
                     /label=chimeric intron
                     /note="chimera between introns from chicken beta-actin and 
                     rabbit beta-globin"
     primer_bind     1606..1628
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1694..1713
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     complement(3480..3499)
                     /label=Bglob-pA-R
                     /note="Rabbit beta-globin polyA region, reverse primer"
     polyA_signal    3545..3600
                     /label=beta-globin poly(A) signal
                     /note="rabbit beta-globin polyadenylation signal (Gil and 
                     Proudfoot, 1987)"
     primer_bind     complement(3599..3618)
                     /label=rbglobpA-R
                     /note="Rabbit beta-globin polyA, reverse primer. Also 
                     called rb-glob-pA-term-R"
     primer_bind     complement(3961..3977)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(3961..3977)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(3974..3996)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    3985..4001
                     /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(4009..4015,4016..4033,4034..4039))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4054..4075
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        4134..4329
                     /label=SV40 promoter
                     /note="SV40 early promoter"
     rep_origin      4180..4315
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     polyA_signal    4335..4469
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4372..4391)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4426..4445
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(4537..4554)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(4708..5296)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4788..4807)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5467..6258,6259..6327))
                     /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     6090..6109
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6328..6432)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 ggtcgatcga cattgattat tgactagtta ttaatagtaa tcaattacgg ggtcattagt
       61 tcatagccca tatatggagt tccgcgttac ataacttacg gtaaatggcc cgcctggctg
      121 accgcccaac gacccccgcc cattgacgtc aataatgacg tatgttccca tagtaacgcc
      181 aatagggact ttccattgac gtcaatgggt ggagtattta cggtaaactg cccacttggc
      241 agtacatcaa gtgtatcata tgccaagtac gccccctatt gacgtcaatg acggtaaatg
      301 gcccgcctgg cattatgccc agtacatgac cttatgggac tttcctactt ggcagtacat
      361 ctacgtatta gtcatcgcta ttaccatggt cgaggtgagc cccacgttct gcttcactct
      421 ccccatctcc cccccctccc cacccccaat tttgtattta tttatttttt aattattttg
      481 tgcagcgatg ggggcggggg gggggggggg gcgcgccrgg gggggggggg gggggggggg
      541 gggggggsgg ggsgrggcgg agaggtgcgg cggcagccaa tcagagcggc gcgctccgaa
      601 agtttccttt tatggcgagg cggcggcggc ggcggcccta taaaaagcga agcgcgcggc
      661 gggcgggagt cgctgcgcgc tgccttcgcc ccgtgccccg ctccgccgcc gcctcgcgcc
      721 gcccgccccg gctctgactg accgcgttac tcccacaggt gagcgggcgg gacggccctt
      781 ctcctccggg ctgtaattag cgcttggttt aatgacggct tgtttctttt ctgtggctgc
      841 gtgaaagcct tgaggggctc cgggagggcc ctttgtgcgg ggggagcggc tcggggggtg
      901 cgtgcgtgtg tgtgtgcgtg gggagcgccg cgtgcggctc cgcgctgccc ggcggctgtg
      961 agcgctgcgg gcgcggcgcg gggctttgtg cgctccgcag tgtgcgcgag gggagcgcgg
     1021 ccgggggcgg tgccccgcgg tgcggggggg gctgcgaggg gaacaaaggc tgcgtgcggg
     1081 gtgtgtgcgt gggggggtga gcagggggtg tgggcgcgtc ggtcgggctg caaccccccc
     1141 tgcacccccc tccccgagtt gctgagcacg gcccggcttc gggtgcgggg ctccgtacgg
     1201 ggcgtggcgc ggggctcgcc gtgccgggcg gggggtggcg gcaggtgggg gtgccgggcg
     1261 gggcggggcc gcctcgggcc ggggagggct cgggggaggg gcgcggcggc ccccggagcg
     1321 ccggcggctg tcgaggcgcg gcgagccgca gccattgcct tttatggtaa tcgtgcgaga
     1381 gggcgcaggg acttcctttg tcccaaatct gtgcggagcc gaaatctggg aggcgccgcc
     1441 gcaccccctc tagcgggcgc ggggcgaagc ggtgcggcgc cggcaggaag gaaatgggcg
     1501 gggagggcct tcgtgcgtcg ccgcgccgcc gtccccttct ccctctccag cctcggggct
     1561 gtccgcgggg ggacggctgc cttcgggggg gacggggcag ggcggggttc ggcttctggc
     1621 gtgtgaccgg cggctctagc tagagcctct gctaaccatg ttcatgcctt cttctttttc
     1681 ctacagctcc tgggcaacgt gctggttatt gtgctgtctc atcattttgg caagaattca
     1741 ccatggtgag caagggcgag gagaccacaa tgggcgtaat caagcccgac atgaagatca
     1801 agctgaagat ggagggcaac gtgaatggcc acgccttcgt gatcgagggc gagggcgagg
     1861 gcaagcccta cgacggcacc aacaccatca acctggaggt gaaggaggga gcccccctgc
     1921 ccttctccta cgacattctg accaccgcgt tcgcctacgg caacagggcc ttcaccaagt
     1981 accccgacga catccccaac tacttcaagc agtccttccc cgagggctac tcttgggagc
     2041 gcaccatgac cttcgaggac aagggcatcg tgaaggtgaa gtccgacatc tccatggagg
     2101 aggactcctt catctacgag atacacctca agggcgagaa cttccccccc aacggccccg
     2161 tgatgcagaa gaagaccacc ggctgggacg cctccaccga gaggatgtac gtgcgcgacg
     2221 gcgtgctgaa gggcgacgtc aagcacaagc tgctgctgga gggcggcggc caccaccgcg
     2281 ttgacttcaa gaccatctac agggccaaga aggcggtgaa gctgcccgac tatcactttg
     2341 tggaccaccg catcgagatc ctgaaccacg acaaggacta caacaaggtg accgtttacg
     2401 agagcgccgt ggcccgcatg atgaagcggc tggtgtacat cagcaagatc agcggccacc
     2461 tgagcctgga agagatccag cggatcggca aggtgtccat caagaacaac cagcgggaca
     2521 acatcaccgg cgtgctgctg tacctgcagg gcctgttctt ccagatcctg gaaggcgaga
     2581 acgagaaggt ggacaagctg tacaagaaaa tcctggtgga cgaccggcac accaacatcc
     2641 tgtgcctgaa aaccgagtac gacatcaccg accggatgtt ccccaactgg gccatgaaaa
     2701 ccatcaacct gaacgagaac agcgagctga tgatccagcc catcaagagc ctgctgcaga
     2761 ccatcaccca gagccaccgg gtgctggaaa agtacatgcc cgccagagtg atctacctga
     2821 tcaaccaggg catcaacccc ctgaccgtgg agccccagct ggtcgagaag atcatcttct
     2881 tcagcgacat cctggccttc agcaccctga ccgagaagct gcccgtgaac gaggtggtca
     2941 tcctggtcaa ccggtacttc agcatctgca cccggatcat cagcgcctac ggcggcgaag
     3001 tgaccaagtt catcggcgac tgcgtgatgg ccagcttcac caaagagcag ggcgacgccg
     3061 ccatccggac cagcctggac atcatctccg agctgaagca gctgcggcac cacgtggagg
     3121 ccaccaaccc cctgcacctg ctgtacaccg gcatcggcct gagctacggc cacgtgatcg
     3181 agggcaacat gggcagcagc ctgaagatgg accacaccct gctgggggac gccgtgaacg
     3241 tggccgccag gctggaagcc ctgacaagac agctgcccta cgccctggcc tttacagccg
     3301 gcgtgaagaa gtgctgccag gcccagtgga ccttcatcaa cctgggagcc caccaggtca
     3361 agggcaagca ggaagccatc gaggtctaca ccgtgaatga ggcccagaag tactacgaca
     3421 ccctgcagat cacccagctg atccggcaga ccctggaaaa cgacaagtaa gtcgacggat
     3481 ctttttccct ctgccaaaaa ttatggggac atcatgaagc cccttgagca tctgacttct
     3541 ggctaataaa ggaaatttat tttcattgca atagtgtgtt ggaatttttt gtgtctctca
     3601 ctcggaagga catatgggag ggcaaatcat ttaaaacatc agaatgagta tttggtttag
     3661 agtttggcaa catatgccca tatgctggct gccatgaaca aaggttggct ataaagaggt
     3721 catcagtata tgaaacagcc ccctgctgtc cattccttat tccatagaaa agccttgact
     3781 tgaggttaga ttttttttat attttgtttt gtgttatttt tttctttaac atccctaaaa
     3841 ttttccttac atgttttact agccagattt ttcctcctct cctgactact cccagtcata
     3901 gctgtccctc ttctcttatg gagatccctc gacctgcagc ccaagcttgg cgtaatcatg
     3961 gtcatagctg tttcctgtgt gaaattgtta tccgctcaca attccacaca acatacgagc
     4021 cggaagcata aagtgtaaag cctggggtgc ctaatgagtg agctaactca cattaattgc
     4081 gttgcgctca ctgcccgctt tccagtcggg aaacctgtcg tgccagcgga tccgcatctc
     4141 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     4201 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     4261 gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     4321 ttttgcaaaa agctaacttg tttattgcag cttataatgg ttacaaataa agcaatagca
     4381 tcacaaattt cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac
     4441 tcatcaatgt atcttatcat gtctggatcc gctgcattaa tgaatcggcc aacgcgcggg
     4501 gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact cgctgcgctc
     4561 ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac ggttatccac
     4621 agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa aggccaggaa
     4681 ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg acgagcatca
     4741 caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa gataccaggc
     4801 gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc ttaccggata
     4861 cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac gctgtaggta
     4921 tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac cccccgttca
     4981 gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg taagacacga
     5041 cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt atgtaggcgg
     5101 tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa cagtatttgg
     5161 tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct cttgatccgg
     5221 caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga ttacgcgcag
     5281 aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg ctcagtggaa
     5341 cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct tcacctagat
     5401 ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt aaacttggtc
     5461 tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc tatttcgttc
     5521 atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg gcttaccatc
     5581 tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag atttatcagc
     5641 aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt tatccgcctc
     5701 catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag ttaatagttt
     5761 gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt ttggtatggc
     5821 ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca tgttgtgcaa
     5881 aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg ccgcagtgtt
     5941 atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat ccgtaagatg
     6001 cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta tgcggcgacc
     6061 gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca gaactttaaa
     6121 agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct taccgctgtt
     6181 gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat cttttacttt
     6241 caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa agggaataag
     6301 ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt gaagcattta
     6361 tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa ataaacaaat
     6421 aggggttccg cgcacatttc cccgaaaagt gccacctg
//
