LOCUS       sequence-306852-        7033 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  For packaging and expression of Chronos-mT-Sapphire featuring a 
            neuritin (NRN) 3' UTR in neurons via AAV.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7033)
  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 7033)
  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-NRN
FEATURES             Location/Qualifiers
     source          1..7033
                     /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        225..672
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      685..694
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1681..1683,1684..2394)
                     /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(1723..1744)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1984..2003)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2331..2352
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    2429..3017
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2482..2502)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2900..2911)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4043..4060)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4049..4256
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4296..4436
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4296..4425
                     /label=AAV2 ITR
     rep_origin      4511..4966
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4598..4617)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4808..4829
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4983..5002)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5102..5124
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5162..5180)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5248..5352
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5353..5421,5422..6213)
                     /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(5571..5590)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6384..6972
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6873..6892
                     /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 gcggccgcac gcgtttaaca ttatggcctt aggtcacttc atctccatgg
      181 ggttcttctt ctgattttct agactgcaga gggccctgcg tatgagtgca agtgggtttt
      241 aggaccagga tgaggcgggg tgggggtgcc tacctgacga ccgaccccga cccactggac
      301 aagcacccaa cccccattcc ccaaattgcg catcccctat cagagagggg gaggggaaac
      361 aggatgcggc gaggcgcgtg cgcactgcca gcttcagcac cgcggacagt gccttcgccc
      421 ccgcctggcg gcgcgcgcca ccgccgcctc agcactgaag gcgcgctgac gtcactcgcc
      481 ggtcccccgc aaactcccct tcccggccac cttggtcgcg tccgcgccgc cgccggccca
      541 gccggaccgc accacgcgag gcgcgagata ggggggcacg ggcgcgacca tctgcgctgc
      601 ggcgccggcg actcagcgct gcctcagtct gcggtgggca gcggaggagt cgtgtcgtgc
      661 ctgagagcgc agtcgagagg atccgccacc atggaaacag ccgccacaat gacccacgcc
      721 tttatctcag ccgtgcctag cgccgaagcc acaattagag gcctgctgag cgccgcagca
      781 gtggtgacac cagcagcaga cgctcacgga gaaacctcta acgccacaac agccggagcc
      841 gatcacggtt gcttccccca catcaaccac ggaaccgagc tgcagcacaa gatcgcagtg
      901 ggactccagt ggttcaccgt gatcgtggct atcgtgcagc tcatcttcta cggttggcac
      961 agcttcaagg ccacaaccgg ctgggaggag gtctacgtct gcgtgatcga gctcgtcaag
     1021 tgcttcatcg agctgttcca cgaggtcgac agcccagcca cagtgtacca gaccaacgga
     1081 ggagccgtga tttggctgcg gtacagcatg tggctcctga cttgccccgt gatcctgatc
     1141 cacctgagca acctgaccgg actgcacgaa gagtacagca agcggaccat gaccatcctg
     1201 gtgaccgaca tcggcaacat cgtgtggggg atcacagccg cctttacaaa gggccccctg
     1261 aagatcctgt tcttcatgat cggcctgttc tacggcgtga cttgcttctt ccagatcgcc
     1321 aaggtgtata tcgagagcta ccacaccctg cccaaaggcg tctgccggaa gatttgcaag
     1381 atcatggcct acgtcttctt ctgctcttgg ctgatgttcc ccgtgatgtt catcgccgga
     1441 cacgagggac tgggcctgat cacaccttac accagcggaa tcggccacct gatcctggat
     1501 ctgatcagca agaacacttg gggcttcctg ggccaccacc tgagagtgaa gatccacgag
     1561 cacatcctga tccacggcga catccggaag acaaccacca tcaacgtggc cggcgagaac
     1621 atggagatcg agaccttcgt cgacgaggag gagggaggag tggcggcacc ggtagtagca
     1681 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     1741 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
     1801 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     1861 gtgaccacct tctcctacgg cgtgatggtg ttcgcccgct accccgacca catgaagcag
     1921 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
     1981 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     2041 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     2101 ctggagtaca acttcaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc
     2161 atcaaggcca acttcaagat ccgccacaac atcgaggacg gcggcgtgca gctcgccgac
     2221 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     2281 ctgagcatcc agtccaagct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     2341 ctggagttcg tgaccgccgc cgggatcact ctgggcatgg acgagctgta caagtccgga
     2401 ctcagatctc gagctcaagc ttatcgataa tcaacctctg gattacaaaa tttgtgaaag
     2461 attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat
     2521 gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc
     2581 ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg
     2641 cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct
     2701 ttccggaact ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct
     2761 tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg
     2821 gaaatcatcg tcctttcctt ggctgctcgc ctatgttgcc acctggattc tgcgcgggac
     2881 gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct
     2941 gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct
     3001 ttgggccgcc tccccgcatc gataccgagc gctgctcgag agatctgcgt ggggccagct
     3061 ccccccgcgc gcccacccac actcactcca tgctcccgga aatcgagagg aagatccatt
     3121 agttctttgg ggacgttgtg attctctgtg atgctgaaaa cactcatata ggattgtggg
     3181 aaatcctgat tctctttttt atttcgtttg atttcttgtg ttttatttgc caaatgttac
     3241 caatcagtga gcaagcaagc acagccaaaa tcggacctca gctttagtcc gtcttcacac
     3301 acaaataaga aaacggcaaa cccaccccat tttttaattt tattattatt aatttttttt
     3361 gttggcaaaa gaatctcagg aacggccctg ggccacctac tatattaatc atgctagtaa
     3421 catgaaaaat gatgggctcc tcctaatagg aaggcgagga gaggagaagg ccaggggaat
     3481 gaattcaaga gagatgtcca cggacgaaac atacggtgaa taattcacgc tcacgtcgtt
     3541 cttccacagt atcttgtttt gatcatttcc actgcacatt tctcctcaag aaaagcgaaa
     3601 ggacagactg ttggctttgt gtttggagga taggagggag agagggaagg ggctgaggaa
     3661 atctctgggg taagagtaaa ggcttccaga agacatgctg ctatggtcac tgaggggtta
     3721 gctttatctg ctgttgttga tgcatccgtc caagttcact gcctttattt tccctcctcc
     3781 ctcttgtttt agctgttaca cacacagtaa tacctgaata tccaacggta tagatcacaa
     3841 gggggggatg ttaaatgtta atctaaaata tagctaaaaa aagattttga cataaaagag
     3901 ccttgatttt aaaaaaaaaa gagagagaga tgtaatttaa aaagtttatt ataaattaaa
     3961 ttcagcaaaa aaagatttgc tacaaagtat agagaagtat aaaataaaag ttattgtttg
     4021 aaaaaaaaaa aaaaaaaata agcctcgact gtgccttcta gttgccagcc atctgttgtt
     4081 tgcccctccc ccgtgccttc cttgaccctg gaaggtgcca ctcccactgt cctttcctaa
     4141 taaaatgagg aaattgcatc gcattgtctg agtaggtgtc attctattct ggggggtggg
     4201 gtggggcagg acagcaaggg ggaggattgg gaagacaata gcaggcatgc tggggactga
     4261 ttttgtaggt aaccacgtgc ggaccgagcg gccgcaggaa cccctagtga tggagttggc
     4321 cactccctct ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg
     4381 cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg
     4441 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag
     4501 caaccatagt acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     4561 agcgtgaccg ctacacttgc cagcgcctta gcgcccgctc ctttcgcttt cttcccttcc
     4621 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     4681 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca
     4741 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     4801 tttaatagtg gactcttgtt ccaaactgga acaacactca actctatctc gggctattct
     4861 tttgatttat aagggatttt gccgatttcg gtctattggt taaaaaatga gctgatttaa
     4921 caaaaattta acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc
     4981 agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct
     5041 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     5101 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag
     5161 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     5221 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     5281 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     5341 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     5401 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     5461 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     5521 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     5581 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     5641 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     5701 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     5761 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     5821 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     5881 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     5941 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     6001 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     6061 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     6121 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     6181 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     6241 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     6301 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     6361 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     6421 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     6481 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     6541 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     6601 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     6661 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     6721 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     6781 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     6841 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     6901 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     6961 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     7021 tttgctcaca tgt
//
