LOCUS       sequence-478672-        5275 bp    DNA     circular SYN 15-JUL-2025
DEFINITION  Cre-dependent, synapsin promoter-driven expression of the green 
            fluorescent ExRai-AKAR2 PKA biosensor in neurons..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5275)
  AUTHORS   Zhang JF, Liu B, Hong I, Mo A, Roth RH, Tenner B, Lin W, Zhang JZ, 
            Molina RS, Drobizhev M, Hughes TE, Tian L, Huganir RL, Mehta S, 
            Zhang J
  TITLE     An ultrasensitive biosensor for high-resolution kinase activity 
            imaging in awake mice.
  JOURNAL   Nat Chem Biol. 2020 Sep 28. pii: 10.1038/s41589-020-00660-y. doi: 
            10.1038/s41589-020-00660-y.
  PUBMED    32989297
REFERENCE   2  (bases 1 to 5275)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jul 15, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "Nat Chem 
            Biol. 2020 Sep 28. pii: 10.1038/s41589-020-00660-y. doi: 
            10.1038/s41589-020-00660-y."
COMMENT     Sequence Label: pAAV-hSyn1-DIO-ExRai-AKAR2
FEATURES             Location/Qualifiers
     source          1..5275
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        159..606
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    complement(607..640)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(671..704)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     1319..1338
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1578..1599
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1705..1726)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    2004..2037
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    2068..2101
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     primer_bind     complement(2158..2178)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     repeat_region   2538..2678
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2538..2667
                     /label=AAV2 ITR
     rep_origin      2753..3208
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2840..2859)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3050..3071
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3225..3244)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3344..3366
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3404..3422)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3490..3594
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3595..3663,3664..4455)
                     /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     complement(3813..3832)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4626..5214
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5115..5134
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgctg cagagggccc tgcgtatgag tgcaagtggg ttttaggacc
      181 aggatgaggc ggggtggggg tgcctacctg acgaccgacc ccgacccact ggacaagcac
      241 ccaaccccca ttccccaaat tgcgcatccc ctatcagaga gggggagggg aaacaggatg
      301 cggcgaggcg cgtgcgcact gccagcttca gcaccgcgga cagtgccttc gcccccgcct
      361 ggcggcgcgc gccaccgccg cctcagcact gaaggcgcgc tgacgtcact cgccggtccc
      421 ccgcaaactc cccttcccgg ccaccttggt cgcgtccgcg ccgccgccgg cccagccgga
      481 ccgcaccacg cgaggcgcga gatagggggg cacgggcgcg accatctgcg ctgcggcgcc
      541 ggcgactcag cgctgcctca gtctgcggtg ggcagcggag gagtcgtgtc gtgcctgaga
      601 gcgcagataa cttcgtatag gatactttat acgaagttat gcagaatggt agctggattg
      661 tagctgctat ataacttcgt atagcataca ttatacgaag ttatctaaat tttaacactc
      721 tcgatgccat atacgttgta accttcctta taagttgcaa aattctggga attcctagcg
      781 atcaactttg ttctgctcga ggcactgctt aaatttgtcg tttatgaaaa tgaccagaga
      841 taaaatatct gattccacgc ctacaccaac ggttatttca tcaccttgag acagtaactg
      901 attgctgttc ttctcgactt tttgcccatt taaccaggtc ccattagtgg aaatgtcatt
      961 caataaaagg ttaccgtctt ctcctagtag tatttggaaa tgcttatttg acagtcttga
     1021 aatgtttcct aaatgatagt cacaggctgg gtttctacca aatgtccaaa ctttctttat
     1081 ggatcgtttt tccttaagca cttgtgaaat atcagctgac aaatctcgga tgggaatttg
     1141 acccgtggta caaatgaccc tgcacacaat gttttcgccg atctgttctt gagaaaactt
     1201 atgcagcaag ttgtactcca gcttgtgccc caggatgttg ccgtcctcct tgaagtcgat
     1261 gcccttcagc tcgatgcggt tcaccagggt gtcgccctcg aacttcacct cggcgcgggt
     1321 cttgtagttg ccgtcgtcct tgaagaagat ggtgcgctcc tggatgtagc cttcgggcat
     1381 ggcggacttg aagaagtcgt gctgcttcat gtggtcgggg tagcggctga agcactgcac
     1441 gccgtaggtc agggtggtca cgagggtggg ccagggcacg ggcagcttgc cggtggtgca
     1501 gatgaacttc agggtcagct tgccgtaggt ggcatcgccc tcaccctcgc cggacacgct
     1561 gaacttgtgg ccgtttacgt cgccgtccag ctcgaccagg atgggcacca ccccggtgaa
     1621 cagctcctcg cccttgctca ccatgctccc tccggtaccg cccttgtaca gctcgtccat
     1681 gccgagagtg atcccggcgg cggtcacgaa ctccagcagg accatgtgat cgcgcttctc
     1741 gttggggtct ttcgaaagtt tggactgcac gctcaggtag tggttgtcgg gcagcagcac
     1801 ggggccgtcg ccgatggggg tgttctgctg gtagtggtag gcgagctgca cgccgccgtc
     1861 ctcgatgttg tggcggatct tgaagttcgc cttgatgccg ttcttctgct tgtcggcctt
     1921 gatatagacg ttgcagaagc tgagctcgct gccgccggtg ccgccgtcca ccagggtggc
     1981 gcgacgcagc atggtggctc gagataactt cgtataaagt atcctatacg aagttatttg
     2041 ccttaaccca gaaattatca ctgttatata acttcgtata atgtatgcta tacgaagtta
     2101 tgataatcaa cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt
     2161 tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc
     2221 ccgtatggct ttcattttct cctccttgta taaatcctgg ttagttcttg ccacggcgga
     2281 actcatcgcc gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa
     2341 ttccgtggtg tttatttgtg aaatttgtga tgctattgct ttatttgtaa ccatctagct
     2401 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac
     2461 aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag atgtgggagg
     2521 ttttttaaag cggccgcagg aacccctagt gatggagttg gccactccct ctctgcgcgc
     2581 tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc
     2641 ggcctcagtg agcgagcgag cgcgcagctg cctgcagggg cgcctgatgc ggtattttct
     2701 ccttacgcat ctgtgcggta tttcacaccg catacgtcaa agcaaccata gtacgcgccc
     2761 tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac cgctacactt
     2821 gccagcgcct tagcgcccgc tcctttcgct ttcttccctt cctttctcgc cacgttcgcc
     2881 ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt tagtgcttta
     2941 cggcacctcg accccaaaaa acttgatttg ggtgatggtt cacgtagtgg gccatcgccc
     3001 tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag tggactcttg
     3061 ttccaaactg gaacaacact caactctatc tcgggctatt cttttgattt ataagggatt
     3121 ttgccgattt cggtctattg gttaaaaaat gagctgattt aacaaaaatt taacgcgaat
     3181 tttaacaaaa tattaacgtt tacaatttta tggtgcactc tcagtacaat ctgctctgat
     3241 gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc ctgacgggct
     3301 tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag ctgcatgtgt
     3361 cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt gatacgccta
     3421 tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg cacttttcgg
     3481 ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa tatgtatccg
     3541 ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa gagtatgagt
     3601 attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct tcctgttttt
     3661 gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg tgcacgagtg
     3721 ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg ccccgaagaa
     3781 cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt atcccgtatt
     3841 gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga cttggttgag
     3901 tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga attatgcagt
     3961 gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac gatcggagga
     4021 ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg ccttgatcgt
     4081 tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac gatgcctgta
     4141 gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct agcttcccgg
     4201 caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct gcgctcggcc
     4261 cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg gtctcgcggt
     4321 atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat ctacacgacg
     4381 gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg tgcctcactg
     4441 attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat tgatttaaaa
     4501 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     4561 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     4621 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     4681 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     4741 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac
     4801 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     4861 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     4921 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     4981 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     5041 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     5101 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     5161 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     5221 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgt
//
