LOCUS       sequence-306850-        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"
     repeat_region   1..130
                     /label=AAV2 ITR
     promoter        230..677
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      690..699
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1686..1688,1689..2399)
                     /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(1728..1749)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1989..2008)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2336..2357
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    2434..3022
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2487..2507)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2905..2916)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3063..3270
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   3440..3569
                     /label=AAV2 ITR
     rep_origin      complement(3806..4394)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3886..3905)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4565..5356,5357..5425))
                     /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     5188..5207
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5426..5530)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5556..6011)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5693..5714)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5905..5924
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct
      181 aggaagatcg taggttaatt aatctagact gcagagggcc ctgcgtatga gtgcaagtgg
      241 gttttaggac caggatgagg cggggtgggg gtgcctacct gacgaccgac cccgacccac
      301 tggacaagca cccaaccccc attccccaaa ttgcgcatcc cctatcagag agggggaggg
      361 gaaacaggat gcggcgaggc gcgtgcgcac tgccagcttc agcaccgcgg acagtgcctt
      421 cgcccccgcc tggcggcgcg cgccaccgcc gcctcagcac tgaaggcgcg ctgacgtcac
      481 tcgccggtcc cccgcaaact ccccttcccg gccaccttgg tcgcgtccgc gccgccgccg
      541 gcccagccgg accgcaccac gcgaggcgcg agataggggg gcacgggcgc gaccatctgc
      601 gctgcggcgc cggcgactca gcgctgcctc agtctgcggt gggcagcgga ggagtcgtgt
      661 cgtgcctgag agcgcagtcg agaggatccg ccaccatgga aacagccgcc acaatgaccc
      721 acgcctttat ctcagccgtg cctagcgccg aagccacaat tagaggcctg ctgagcgccg
      781 cagcagtggt gacaccagca gcagacgctc acggagaaac ctctaacgcc acaacagccg
      841 gagccgatca cggttgcttc ccccacatca accacggaac cgagctgcag cacaagatcg
      901 cagtgggact ccagtggttc accgtgatcg tggctatcgt gcagctcatc ttctacggtt
      961 ggcacagctt caaggccaca accggctggg aggaggtcta cgtctgcgtg atcgagctcg
     1021 tcaagtgctt catcgagctg ttccacgagg tcgacagccc agccacagtg taccagacca
     1081 acggaggagc cgtgatttgg ctgcggtaca gcatgtggct cctgacttgc cccgtgatcc
     1141 tgatccacct gagcaacctg accggactgc acgaagagta cagcaagcgg accatgacca
     1201 tcctggtgac cgacatcggc aacatcgtgt gggggatcac agccgccttt acaaagggcc
     1261 ccctgaagat cctgttcttc atgatcggcc tgttctacgg cgtgacttgc ttcttccaga
     1321 tcgccaaggt gtatatcgag agctaccaca ccctgcccaa aggcgtctgc cggaagattt
     1381 gcaagatcat ggcctacgtc ttcttctgct cttggctgat gttccccgtg atgttcatcg
     1441 ccggacacga gggactgggc ctgatcacac cttacaccag cggaatcggc cacctgatcc
     1501 tggatctgat cagcaagaac acttggggct tcctgggcca ccacctgaga gtgaagatcc
     1561 acgagcacat cctgatccac ggcgacatcc ggaagacaac caccatcaac gtggccggcg
     1621 agaacatgga gatcgagacc ttcgtcgacg aggaggaggg aggagtggcg gcaccggtag
     1681 tagcagtgag caagggcgag gagctgttca ccggggtggt gcccatcctg gtcgagctgg
     1741 acggcgacgt aaacggccac aagttcagcg tgtccggcga gggcgagggc gatgccacct
     1801 acggcaagct gaccctgaag ttcatctgca ccaccggcaa gctgcccgtg ccctggccca
     1861 ccctcgtgac caccttctcc tacggcgtga tggtgttcgc ccgctacccc gaccacatga
     1921 agcagcacga cttcttcaag tccgccatgc ccgaaggcta cgtccaggag cgcaccatct
     1981 tcttcaagga cgacggcaac tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc
     2041 tggtgaaccg catcgagctg aagggcatcg acttcaagga ggacggcaac atcctggggc
     2101 acaagctgga gtacaacttc aacagccaca acgtctatat catggccgac aagcagaaga
     2161 acggcatcaa ggccaacttc aagatccgcc acaacatcga ggacggcggc gtgcagctcg
     2221 ccgaccacta ccagcagaac acccccatcg gcgacggccc cgtgctgctg cccgacaacc
     2281 actacctgag catccagtcc aagctgagca aagaccccaa cgagaagcgc gatcacatgg
     2341 tcctgctgga gttcgtgacc gccgccggga tcactctggg catggacgag ctgtacaagt
     2401 ccggactcag atctcgagct caagcttatc gataatcaac ctctggatta caaaatttgt
     2461 gaaagattga ctggtattct taactatgtt gctcctttta cgctatgtgg atacgctgct
     2521 ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt tcattttctc ctccttgtat
     2581 aaatcctggt tgctgtctct ttatgaggag ttgtggcccg ttgtcaggca acgtggcgtg
     2641 gtgtgcactg tgtttgctga cgcaaccccc actggttggg gcattgccac cacctgtcag
     2701 ctcctttccg ggactttcgc tttccccctc cctattgcca cggcggaact catcgccgcc
     2761 tgccttgccc gctgctggac aggggctcgg ctgttgggca ctgacaattc cgtggtgttg
     2821 tcggggaaat catcgtcctt tccttggctg ctcgcctgtg ttgccacctg gattctgcgc
     2881 gggacgtcct tctgctacgt cccttcggcc ctcaatccag cggaccttcc ttcccgcggc
     2941 ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc gccctcagac gagtcggatc
     3001 tccctttggg ccgcctcccc gcatcgatac cgtcgacccg ggcggccgct cataaagctc
     3061 gactgtgcct tctagttgcc agccatctgt tgtttgcccc tcccccgtgc cttccttgac
     3121 cctggaaggt gccactccca ctgtcctttc ctaataaaat gaggaaattg catcgcattg
     3181 tctgagtagg tgtcattcta ttctgggggg tggggtgggg caggacagca agggggagga
     3241 ttgggaagac aatagcaggc atgctgggga tgcggtccgg actgtacaat tgcattcatt
     3301 ttatgtttca ggttcagggg gagatgtggg aggtttttta aagcaagtaa aacctctaca
     3361 aatgtggtaa aatcgataag gatcttccta gagcatggct acgtagataa gtagcatggc
     3421 gggttaatca ttaactacaa ggaaccccta gtgatggagt tggccactcc ctctctgcgc
     3481 gctcgctcgc tcactgaggc cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg
     3541 gcggcctcag tgagcgagcg agcgcgcagc tgcattaatg aatcggccaa cgcgcgggga
     3601 gaggcggttt gcgtattggg cgctcttccg cttcctcgct cactgactcg ctgcgctcgg
     3661 tcgttcggct gcggcgagcg gtatcagctc actcaaaggc ggtaatacgg ttatccacag
     3721 aatcagggga taacgcagga aagaacatgt gagcaaaagg ccagcaaaag gccaggaacc
     3781 gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg cccccctgac gagcatcaca
     3841 aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg actataaaga taccaggcgt
     3901 ttccccctgg aagctccctc gtgcgctctc ctgttccgac cctgccgctt accggatacc
     3961 tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca tagctcacgc tgtaggtatc
     4021 tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt gcacgaaccc cccgttcagc
     4081 ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc caacccggta agacacgact
     4141 tatcgccact ggcagcagcc actggtaaca ggattagcag agcgaggtat gtaggcggtg
     4201 ctacagagtt cttgaagtgg tggcctaact acggctacac tagaagaaca gtatttggta
     4261 tctgcgctct gctgaagcca gttaccttcg gaaaaagagt tggtagctct tgatccggca
     4321 aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa gcagcagatt acgcgcagaa
     4381 aaaaaggatc tcaagaagat cctttgatct tttctacggg gtctgacgct cagtggaacg
     4441 aaaactcacg ttaagggatt ttggtcatga gattatcaaa aaggatcttc acctagatcc
     4501 ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat atatgagtaa acttggtctg
     4561 acagttacca atgcttaatc agtgaggcac ctatctcagc gatctgtcta tttcgttcat
     4621 ccatagttgc ctgactcccc gtcgtgtaga taactacgat acgggagggc ttaccatctg
     4681 gccccagtgc tgcaatgata ccgcgagacc cacgctcacc ggctccagat ttatcagcaa
     4741 taaaccagcc agccggaagg gccgagcgca gaagtggtcc tgcaacttta tccgcctcca
     4801 tccagtctat taattgttgc cgggaagcta gagtaagtag ttcgccagtt aatagtttgc
     4861 gcaacgttgt tgccattgct acaggcatcg tggtgtcacg ctcgtcgttt ggtatggctt
     4921 cattcagctc cggttcccaa cgatcaaggc gagttacatg atcccccatg ttgtgcaaaa
     4981 aagcggttag ctccttcggt cctccgatcg ttgtcagaag taagttggcc gcagtgttat
     5041 cactcatggt tatggcagca ctgcataatt ctcttactgt catgccatcc gtaagatgct
     5101 tttctgtgac tggtgagtac tcaaccaagt cattctgaga atagtgtatg cggcgaccga
     5161 gttgctcttg cccggcgtca atacgggata ataccgcgcc acatagcaga actttaaaag
     5221 tgctcatcat tggaaaacgt tcttcggggc gaaaactctc aaggatctta ccgctgttga
     5281 gatccagttc gatgtaaccc actcgtgcac ccaactgatc ttcagcatct tttactttca
     5341 ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc cgcaaaaaag ggaataaggg
     5401 cgacacggaa atgttgaata ctcatactct tcctttttca atattattga agcatttatc
     5461 agggttattg tctcatgagc ggatacatat ttgaatgtat ttagaaaaat aaacaaatag
     5521 gggttccgcg cacatttccc cgaaaagtgc cacctaaatt gtaagcgtta atattttgtt
     5581 aaaattcgcg ttaaattttt gttaaatcag ctcatttttt aaccaatagg ccgaaatcgg
     5641 caaaatccct tataaatcaa aagaatagac cgagataggg ttgagtgttg ttccagtttg
     5701 gaacaagagt ccactattaa agaacgtgga ctccaacgtc aaagggcgaa aaaccgtcta
     5761 tcagggcgat ggcccactac gtgaaccatc accctaatca agttttttgg ggtcgaggtg
     5821 ccgtaaagca ctaaatcgga accctaaagg gagcccccga tttagagctt gacggggaaa
     5881 gccggcgaac gtggcgagaa aggaagggaa gaaagcgaaa ggagcgggcg ctagggcgct
     5941 ggcaagtgta gcggtcacgc tgcgcgtaac caccacaccc gccgcgctta atgcgccgct
     6001 acagggcgcg tcccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
     6061 ggcctcttcg ctattacgcc ag
//
