LOCUS       sequence-466644-        6551 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  hSyn1 driven expression of the inactive PKA activity recorder 
            negative control with T/A mutation Kinprola_PKA fused to mEGFP for 
            neuronal expression through AAV transduction.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6551)
  AUTHORS   Sun DE, Ng SW, Zheng Y, Xie S, Schwan N, Breuer P, Hoffmann DC, 
            Michel J, Azorin DD, Boonekamp KE, Winkler F, Wick W, Boutros M, Li 
            Y, Johnsson K
  TITLE     Molecular recording of cellular protein kinase activity with 
            chemical labeling.
  JOURNAL   Nat Chem Biol. 2025 Jul 10. doi: 10.1038/s41589-025-01949-6.
  PUBMED    40640555
REFERENCE   2  (bases 1 to 6551)
  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 Chem 
            Biol. 2025 Jul 10. doi: 10.1038/s41589-025-01949-6."
COMMENT     Sequence Label: pAAV_hSyn_NES-Kinprola_PKA_T/A-mEGFP_WRPE-SV40
FEATURES             Location/Qualifiers
     source          1..6551
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     misc_feature    16..604
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(69..89)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(487..498)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(607..623)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(608..624)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    653..774
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(664..683)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     718..737
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   936..1065
                     /label=AAV2 ITR
     rep_origin      complement(1302..1890)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(1382..1401)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2061..2852,2853..2921))
                     /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     2684..2703
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(2922..3026)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(3052..3507)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3189..3210)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3401..3420
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   3579..3708
                     /label=AAV2 ITR
     promoter        3790..4237
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     CDS             join(5835..5837,5838..5840,5841..6551,1..3)
                     /codon_start=1
                     /product="enhanced GFP with monomerizing A206K mutation 
                     (Zacharias et al., 2002)"
                     /label=mEGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(5880..5901)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(6141..6160)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6488..6509
                     /label=EGFP-C
                     /note="EGFP, forward primer"
ORIGIN
        1 tgaaagctta tcgataatca acctctggat tacaaaattt gtgaaagatt gactggtatt
       61 cttaactatg ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat
      121 gctattgctt cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct
      181 ctttatgagg agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct
      241 gacgcaaccc ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc
      301 gctttccccc tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg
      361 acaggggctc ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc
      421 tttccttggc tgctcgcctg tgttgccacc tggattctgc gcgggacgtc cttctgctac
      481 gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg
      541 cctcttccgc gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc
      601 ccgcatcgat accgtcgacc tcgacccggg cggccgcttc gagcagacat gataagatac
      661 attgatgagt ttggacaaac cacaactaga atgcagtgaa aaaaatgctt tatttgtgaa
      721 atttgtgatg ctattgcttt atttgtaacc attataagct gcaataaaca agttaacaac
      781 aacaattgca ttcattttat gtttcaggtt cagggggaga tgtgggaggt tttttaaagc
      841 aagtaaaacc tctacaaatg tggtaaaatc gataaggatc ttcctagagc atggctacgt
      901 agataagtag catggcgggt taatcattaa ctacaaggaa cccctagtga tggagttggc
      961 cactccctct ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg
     1021 cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg cgcagctgca ttaatgaatc
     1081 ggccaacgcg cggggagagg cggtttgcgt attgggcgct cttccgcttc ctcgctcact
     1141 gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat cagctcactc aaaggcggta
     1201 atacggttat ccacagaatc aggggataac gcaggaaaga acatgtgagc aaaaggccag
     1261 caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt ttttccatag gctccgcccc
     1321 cctgacgagc atcacaaaaa tcgacgctca agtcagaggt ggcgaaaccc gacaggacta
     1381 taaagatacc aggcgtttcc ccctggaagc tccctcgtgc gctctcctgt tccgaccctg
     1441 ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa gcgtggcgct ttctcatagc
     1501 tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct ccaagctggg ctgtgtgcac
     1561 gaaccccccg ttcagcccga ccgctgcgcc ttatccggta actatcgtct tgagtccaac
     1621 ccggtaagac acgacttatc gccactggca gcagccactg gtaacaggat tagcagagcg
     1681 aggtatgtag gcggtgctac agagttcttg aagtggtggc ctaactacgg ctacactaga
     1741 agaacagtat ttggtatctg cgctctgctg aagccagtta ccttcggaaa aagagttggt
     1801 agctcttgat ccggcaaaca aaccaccgct ggtagcggtg gtttttttgt ttgcaagcag
     1861 cagattacgc gcagaaaaaa aggatctcaa gaagatcctt tgatcttttc tacggggtct
     1921 gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg tcatgagatt atcaaaaagg
     1981 atcttcacct agatcctttt aaattaaaaa tgaagtttta aatcaatcta aagtatatat
     2041 gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg aggcacctat ctcagcgatc
     2101 tgtctatttc gttcatccat agttgcctga ctccccgtcg tgtagataac tacgatacgg
     2161 gagggcttac catctggccc cagtgctgca atgataccgc gagacccacg ctcaccggct
     2221 ccagatttat cagcaataaa ccagccagcc ggaagggccg agcgcagaag tggtcctgca
     2281 actttatccg cctccatcca gtctattaat tgttgccggg aagctagagt aagtagttcg
     2341 ccagttaata gtttgcgcaa cgttgttgcc attgctacag gcatcgtggt gtcacgctcg
     2401 tcgtttggta tggcttcatt cagctccggt tcccaacgat caaggcgagt tacatgatcc
     2461 cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc cgatcgttgt cagaagtaag
     2521 ttggccgcag tgttatcact catggttatg gcagcactgc ataattctct tactgtcatg
     2581 ccatccgtaa gatgcttttc tgtgactggt gagtactcaa ccaagtcatt ctgagaatag
     2641 tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac gggataatac cgcgccacat
     2701 agcagaactt taaaagtgct catcattgga aaacgttctt cggggcgaaa actctcaagg
     2761 atcttaccgc tgttgagatc cagttcgatg taacccactc gtgcacccaa ctgatcttca
     2821 gcatctttta ctttcaccag cgtttctggg tgagcaaaaa caggaaggca aaatgccgca
     2881 aaaaagggaa taagggcgac acggaaatgt tgaatactca tactcttcct ttttcaatat
     2941 tattgaagca tttatcaggg ttattgtctc atgagcggat acatatttga atgtatttag
     3001 aaaaataaac aaataggggt tccgcgcaca tttccccgaa aagtgccacc taaattgtaa
     3061 gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta aatcagctca ttttttaacc
     3121 aataggccga aatcggcaaa atcccttata aatcaaaaga atagaccgag atagggttga
     3181 gtgttgttcc agtttggaac aagagtccac tattaaagaa cgtggactcc aacgtcaaag
     3241 ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga accatcaccc taatcaagtt
     3301 ttttggggtc gaggtgccgt aaagcactaa atcggaaccc taaagggagc ccccgattta
     3361 gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga agggaagaaa gcgaaaggag
     3421 cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg cgtaaccacc acacccgccg
     3481 cgcttaatgc gccgctacag ggcgcgtccc attcgccatt caggctgcgc aactgttggg
     3541 aagggcgatc ggtgcgggcc tcttcgctat tacgccagct gcgcgctcgc tcgctcactg
     3601 aggccgcccg ggcaaagccc gggcgtcggg cgacctttgg tcgcccggcc tcagtgagcg
     3661 agcgagcgcg cagagaggga gtggccaact ccatcactag gggttccttg tagttaatga
     3721 ttaacccgcc atgctactta tctacgtagc catgctctag gaagatctct gcagagggcc
     3781 ctgcgtatga gtgcaagtgg gttttaggac caggatgagg cggggtgggg gtgcctacct
     3841 gacgaccgac cccgacccac tggacaagca cccaaccccc attccccaaa ttgcgcatcc
     3901 cctatcagag agggggaggg gaaacaggat gcggcgaggc gcgtgcgcac tgccagcttc
     3961 agcaccgcgg acagtgcctt cgcccccgcc tggcggcgcg cgccaccgcc gcctcagcac
     4021 tgaaggcgcg ctgacgtcac tcgccggtcc cccgcaaact ccccttcccg gccaccttgg
     4081 tcgcgtccgc gccgccgccg gcccagccgg accgcaccac gcgaggcgcg agataggggg
     4141 gcacgggcgc gaccatctgc gctgcggcgc cggcgactca gcgctgcctc agtctgcggt
     4201 gggcagcgga ggagtcgtgt cgtgcctgag agcgcagtcg aattcaagct gctagcaagg
     4261 atccacccgc caccatgctg cagaacgagc ttgctcttaa gttggctgga cttgatatta
     4321 acaagactgg aggttctgac gtcggccgca agctgatcat cgatcagaac gtttttatcg
     4381 agggtacgct gccgatgggt gtcgtccgcc cgctgactga agtcgagatg gaccattacc
     4441 gcgagccgtt cctgaatcct gttgaccgcg agccactgtg gcgcttccca aacgagctgc
     4501 caatcgccgg tgagccagcg aacatcgtcg cgctggtcga agaatacatg gactggctgc
     4561 accagtcccc tgtcccgaag ctgctgttct ggggcacccc aggcgttctg atcccaccgg
     4621 ccgaagccgc tcgcctggcc aaaagcctgc ctaactgcaa ggctgtggac atcggcccgg
     4681 gtctgaatct gctgcaagaa gacaacccgg acctgatcgg cagcgagatc gcgcgctggc
     4741 tgtctactct ggagattggt ggcaccggtg gctctggcgg taccggtggc tctggcggtt
     4801 ctatcggtac tggctttcca ttcgaccccc attatgtgga agtcctgggc gagcgcatgc
     4861 actacgtcga tgttggtccg cgcgatggca cccctgtgct gttcctgcac ggtaacccga
     4921 cctcctccta cgtgtggcgc aacatcatcc cgcatgttgc accgacccat cgctgcattg
     4981 ctccagacct gatcggtatg ggcaaatccg acaaaccaga cctgggttat ttcttcgacg
     5041 accacgtccg cttcatggat gccttcatcg aagccctggg tctggaagag gtcgtcctgg
     5101 tcattcacga ctggggctcc gctctgggtt tccactgggc caagcgcaat ccagagcgcg
     5161 tcaaaggtat tgcatttatg gagttcatcc gccctatccc gacctgggac gaatgggctc
     5221 cgggttctct gcgtcgcgcc gccctggtgg accctcctcc tccgccaccg ccaccaccgc
     5281 cgccgccgcc accaccacct cctccgcctc ctccaccacc accgccaccg cctccaccac
     5341 ctggcggcag cgcctgtgac tatcatttag gaaacatttc aagactgtca aataagcatt
     5401 tccaaatact actaggagaa gacggtaacc ttttattgaa tgacatttcc actaatggga
     5461 cctggttaaa tgggcaaaaa gtcgagaaga acagctatca gttactgtct caaggtgatg
     5521 aaataaccgt tggtgtaggc gtggaatcag atattttatc tctggtcatt ttcataaacg
     5581 acaaatttaa gcagtgcctc gagcagaaca aagttgatcg cggtggcggt atgcataagt
     5641 tttctcaaga acagatcggc gaaaacattg tgtgcagggt catttgtacc acgggtcaaa
     5701 ttcccatccg agatttgtca gctgatattt cacaagtgct taaggaaaaa cgatccataa
     5761 agaaagtttg gacatttggt agaaacccag gttctgccaa ggagaccttc caggccttcc
     5821 gcagcggagg ttctatggtg agcaagggcg aggagctgtt caccggggtg gtgcccatcc
     5881 tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc gagggcgagg
     5941 gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc aagctgcccg
     6001 tgccctggcc caccctcgtg accaccctga cctacggcgt gcagtgcttc agccgctacc
     6061 ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc tacgtccagg
     6121 agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag gtgaagttcg
     6181 agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag gaggacggca
     6241 acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat atcatggccg
     6301 acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc gaggacggca
     6361 gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc cccgtgctgc
     6421 tgcccgacaa ccactacctg agcacccagt ccaaactgag caaagacccc aacgagaagc
     6481 gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc ggcatggacg
     6541 agctgtacaa g
//
