LOCUS       sequence-292571-        6082 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For packaging and expression of Chronos-mT-Sapphire in neurons via 
            AAV.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6082)
  AUTHORS   Christoffel DJ, Walsh JJ, Hoerbelt P, Heifets BD, Llorach P, Lopez 
            RC, Ramakrishnan C, Deisseroth K, Malenka RC
  TITLE     Selective filtering of excitatory inputs to nucleus accumbens by 
            dopamine and serotonin.
  JOURNAL   Proc Natl Acad Sci U S A. 2021 Jun 15;118(24). pii: 2106648118. doi:
            10.1073/pnas.2106648118.
  PUBMED    34103400
REFERENCE   2  (bases 1 to 6082)
  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: "2021-06-15"; volume: "118(24). pii: 
            2106648118. doi"; pages: " 10.1073/pnas.2106648118"
COMMENT     Sequence Label: pAAV-hSyn-Chronos-mT-Sapphire
FEATURES             Location/Qualifiers
     source          1..6082
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(182..770)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(262..281)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(941..1732,1733..1801))
                     /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     1564..1583
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1802..1906)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1932..2387)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2069..2090)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2281..2300
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   2459..2588
                     /label=AAV2 ITR
     promoter        2688..3135
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      3148..3157
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4144..4146,4147..4857)
                     /codon_start=1
                     /product="strongly enhanced green fluorescent protein 
                     (Kremers et al., 2007)"
                     /label=SGFP2
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFSYGVMVFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNFNSHNVYIMADKQKNGIKA
                     NFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSIQSKLSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(4186..4207)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4447..4466)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     4794..4815
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    4892..5480
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4945..4965)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5363..5374)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    5521..5728
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   5898..6027
                     /label=AAV2 ITR
ORIGIN
        1 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
       61 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
      121 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
      181 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
      241 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
      301 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
      361 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
      421 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
      481 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
      541 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
      601 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
      661 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
      721 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
      781 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
      841 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
      901 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
      961 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     1021 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     1081 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     1141 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     1201 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     1261 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     1321 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     1381 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     1441 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     1501 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     1561 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     1621 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     1681 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     1741 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     1801 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     1861 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     1921 aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     1981 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     2041 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     2101 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     2161 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     2221 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     2281 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     2341 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc attcgccatt
     2401 caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct
     2461 gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc gggcgtcggg cgacctttgg
     2521 tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact ccatcactag
     2581 gggttccttg tagttaatga ttaacccgcc atgctactta tctacgtagc catgctctag
     2641 gaagatcgta ggttaattaa tctagactgc agagggccct gcgtatgagt gcaagtgggt
     2701 tttaggacca ggatgaggcg gggtgggggt gcctacctga cgaccgaccc cgacccactg
     2761 gacaagcacc caacccccat tccccaaatt gcgcatcccc tatcagagag ggggagggga
     2821 aacaggatgc ggcgaggcgc gtgcgcactg ccagcttcag caccgcggac agtgccttcg
     2881 cccccgcctg gcggcgcgcg ccaccgccgc ctcagcactg aaggcgcgct gacgtcactc
     2941 gccggtcccc cgcaaactcc ccttcccggc caccttggtc gcgtccgcgc cgccgccggc
     3001 ccagccggac cgcaccacgc gaggcgcgag ataggggggc acgggcgcga ccatctgcgc
     3061 tgcggcgccg gcgactcagc gctgcctcag tctgcggtgg gcagcggagg agtcgtgtcg
     3121 tgcctgagag cgcagtcgag aggatccgcc accatggaaa cagccgccac aatgacccac
     3181 gcctttatct cagccgtgcc tagcgccgaa gccacaatta gaggcctgct gagcgccgca
     3241 gcagtggtga caccagcagc agacgctcac ggagaaacct ctaacgccac aacagccgga
     3301 gccgatcacg gttgcttccc ccacatcaac cacggaaccg agctgcagca caagatcgca
     3361 gtgggactcc agtggttcac cgtgatcgtg gctatcgtgc agctcatctt ctacggttgg
     3421 cacagcttca aggccacaac cggctgggag gaggtctacg tctgcgtgat cgagctcgtc
     3481 aagtgcttca tcgagctgtt ccacgaggtc gacagcccag ccacagtgta ccagaccaac
     3541 ggaggagccg tgatttggct gcggtacagc atgtggctcc tgacttgccc cgtgatcctg
     3601 atccacctga gcaacctgac cggactgcac gaagagtaca gcaagcggac catgaccatc
     3661 ctggtgaccg acatcggcaa catcgtgtgg gggatcacag ccgcctttac aaagggcccc
     3721 ctgaagatcc tgttcttcat gatcggcctg ttctacggcg tgacttgctt cttccagatc
     3781 gccaaggtgt atatcgagag ctaccacacc ctgcccaaag gcgtctgccg gaagatttgc
     3841 aagatcatgg cctacgtctt cttctgctct tggctgatgt tccccgtgat gttcatcgcc
     3901 ggacacgagg gactgggcct gatcacacct tacaccagcg gaatcggcca cctgatcctg
     3961 gatctgatca gcaagaacac ttggggcttc ctgggccacc acctgagagt gaagatccac
     4021 gagcacatcc tgatccacgg cgacatccgg aagacaacca ccatcaacgt ggccggcgag
     4081 aacatggaga tcgagacctt cgtcgacgag gaggagggag gagtggcggc accggtagta
     4141 gcagtgagca agggcgagga gctgttcacc ggggtggtgc ccatcctggt cgagctggac
     4201 ggcgacgtaa acggccacaa gttcagcgtg tccggcgagg gcgagggcga tgccacctac
     4261 ggcaagctga ccctgaagtt catctgcacc accggcaagc tgcccgtgcc ctggcccacc
     4321 ctcgtgacca ccttctccta cggcgtgatg gtgttcgccc gctaccccga ccacatgaag
     4381 cagcacgact tcttcaagtc cgccatgccc gaaggctacg tccaggagcg caccatcttc
     4441 ttcaaggacg acggcaacta caagacccgc gccgaggtga agttcgaggg cgacaccctg
     4501 gtgaaccgca tcgagctgaa gggcatcgac ttcaaggagg acggcaacat cctggggcac
     4561 aagctggagt acaacttcaa cagccacaac gtctatatca tggccgacaa gcagaagaac
     4621 ggcatcaagg ccaacttcaa gatccgccac aacatcgagg acggcggcgt gcagctcgcc
     4681 gaccactacc agcagaacac ccccatcggc gacggccccg tgctgctgcc cgacaaccac
     4741 tacctgagca tccagtccaa gctgagcaaa gaccccaacg agaagcgcga tcacatggtc
     4801 ctgctggagt tcgtgaccgc cgccgggatc actctgggca tggacgagct gtacaagtcc
     4861 ggactcagat ctcgagctca agcttatcga taatcaacct ctggattaca aaatttgtga
     4921 aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt
     4981 aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa
     5041 atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt
     5101 gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct
     5161 cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg
     5221 ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc
     5281 ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg
     5341 gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct
     5401 gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc
     5461 cctttgggcc gcctccccgc atcgataccg tcgacccggg cggccgctca taaagctcga
     5521 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     5581 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     5641 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     5701 gggaagacaa tagcaggcat gctggggatg cggtccggac tgtacaattg cattcatttt
     5761 atgtttcagg ttcaggggga gatgtgggag gttttttaaa gcaagtaaaa cctctacaaa
     5821 tgtggtaaaa tcgataagga tcttcctaga gcatggctac gtagataagt agcatggcgg
     5881 gttaatcatt aactacaagg aacccctagt gatggagttg gccactccct ctctgcgcgc
     5941 tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc
     6001 ggcctcagtg agcgagcgag cgcgcagctg cattaatgaa tcggccaacg cgcggggaga
     6061 ggcggtttgc gtattgggcg ct
//
