LOCUS       sequence-441012-        7489 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  CaST-IRES for neuron expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7489)
  AUTHORS   Zhang R, Anguiano M, Aarrestad IK, Lin S, Chandra J, Vadde SS, Olson
            DE, Kim CK
  TITLE     Rapid, biochemical tagging of cellular activity history in vivo.
  JOURNAL   Nat Methods. 2024 Sep;21(9):1725-1735. doi: 
            10.1038/s41592-024-02375-7. Epub 2024 Aug 5.
  PUBMED    39103446
REFERENCE   2  (bases 1 to 7489)
  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"; doi: 
            "10.1038/s41592-024-02375-7"; journalName: "Nat Methods"; date: 
            "2024-09"; volume: "21"; issue: "9"; pages: "1725-1735"
COMMENT     Sequence Label: pAAV-Syn-CD4-sTC-M13-GFP-IRES-CaM-V5-sTN
FEATURES             Location/Qualifiers
     source          1..7489
                     /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        181..628
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      643..652
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     sig_peptide     649..708
                     /codon_start=1
                     /product="leader sequence from mouse immunoglobulin kappa 
                     light chain"
                     /label=Ig-kappa leader
                     /translation="METDTLLLWVLLLWVPGSTG"
     CDS             733..759
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             760..786
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             2311..2367
                     /codon_start=1
                     /product="calmodulin-binding peptide from avian smooth 
                     muscle myosin light chain kinase"
                     /label=calmodulin-binding peptide
                     /translation="RRKWNKTGHAVRAIGRLSS"
     CDS             join(2398..2400,2401..2403,2404..3117)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2398..2400,2401..2403,2404..3114)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2704..2723)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3051..3072
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    3130..3682
                     /label=IRES
                     /note="internal ribosome entry site (IRES) of the 
                     encephalomyocarditis virus (EMCV)"
     primer_bind     complement(3274..3291)
                     /label=IRES reverse
                     /note="IRES internal ribosome entry site, reverse primer. 
                     Also called pCDH-rev"
     primer_bind     3501..3520
                     /label=IRES-F
                     /note="IRES internal ribosome entry site, forward primer"
     regulatory      3691..3700
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4210..4251
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     primer_bind     complement(4510..4527)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    4516..4723
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4752..4892
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4752..4881
                     /label=AAV2 ITR
     rep_origin      4967..5422
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5054..5073)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5264..5285
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5439..5458)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5558..5580
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5618..5636)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5704..5808
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5809..5877,5878..6669)
                     /codon_start=1
                     /gene="bla"
                     /product="beta-lactamase"
                     /label=AmpR
                     /note="confers resistance to ampicillin, carbenicillin, and
                     related antibiotics"
                     /translation="MSIQHFRVALIPFFAAFCLPVFAHPETLVKVKDAEDQLGARVGYI
                     ELDLNSGKILESFRPEERFPMMSTFKVLLCGAVLSRIDAGQEQLGRRIHYSQNDLVEYS
                     PVTEKHLTDGMTVRELCSAAITMSDNTAANLLLTTIGGPKELTAFLHNMGDHVTRLDRW
                     EPELNEAIPNDERDTTMPVAMATTLRKLLTGELLTLASRQQLIDWMEVDKVAGPLLRSA
                     LPAGWFIADKSGAGERGSRGIIAALGPDGKPSRIVVIYTTGSQATMDERNRQIAEIGAS
                     LIKHW"
     rep_origin      6840..7428
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7329..7348
                     /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 gcgttgtaca gtgtctagac tgcagagggc cctgcgtatg
      181 agtgcaagtg ggttttagga ccaggatgag gcggggtggg ggtgcctacc tgacgaccga
      241 ccccgaccca ctggacaagc acccaacccc cattccccaa attgcgcatc ccctatcaga
      301 gagggggagg ggaaacagga tgcggcgagg cgcgtgcgca ctgccagctt cagcaccgcg
      361 gacagtgcct tcgcccccgc ctggcggcgc gcgccaccgc cgcctcagca ctgaaggcgc
      421 gctgacgtca ctcgccggtc ccccgcaaac tccccttccc ggccaccttg gtcgcgtccg
      481 cgccgccgcc ggcccagccg gaccgcacca cgcgaggcgc gagatagggg ggcacgggcg
      541 cgaccatctg cgctgcggcg ccggcgactc agcgctgcct cagtctgcgg tgggcagcgg
      601 aggagtcgtg tcgtgcctga gagcgcagtc gagaagaagc ttgccaccat ggagacagac
      661 acactcctgc tatgggtact gctgctctgg gttccaggtt ccactggtga cggggcccag
      721 ccggccagat cttacccata cgatgtgcca gattacgcct atccctatga cgtccctgat
      781 tacgctctcg acttccagaa ggcctccagc atagtctata agaaagaggg ggaacaggtg
      841 gagttctcct tcccactcgc ctttacagtt gaaaagctga cgggcagtgg cgagctgtgg
      901 tggcaggcgg agagggcttc ctcctccaag tcttggatca cctttgacct gaagaacaag
      961 gaagtgtctg taaaacgggt tacccaggac cctaagctcc agatgggcaa gaagctcccg
     1021 ctccacctca ccctgcccca ggccttgcct cagtatgctg gctctggaaa cctcaccctg
     1081 gcccttgaag cgaaaacagg aaagttgcat caggaagtga acctggtggt gatgagagcc
     1141 actcagctcc agaaaaattt gacctgtgag gtgtggggac ccacctcccc taagctgatg
     1201 ctgagcttga aactggagaa caaggaggca aaggtctcga agcgggagaa ggcggtgtgg
     1261 gtgctgaacc ctgaggcggg gatgtggcag tgtctgctga gtgactcggg acaggtcctg
     1321 ctggaatcca acatcaaggt tctgcccaca tggtccaccc cggtgcagcc aatggccctg
     1381 attgtgctgg ggggcgtcgc cggcctcctg cttttcattg ggctaggcat cttcttctgt
     1441 gtcaggtgcc ggcaccgaag gcgcaagggc tcgggctcga cctcgggctc gggcagcggt
     1501 ggctctagag gttctggtgg cagctctgga ggtggacagc tggacggcgg gagcgtggca
     1561 gtcctgcctg tggtcgactc caccaatcag tacctgctgg atcgaatcgg cgagctgaag
     1621 agtggggatg cttgcattgc agaatatcag caggcaggga gaggaagcag agggaggaaa
     1681 tggttctctc cttttggagc taacctgtac ctgagtatgt tttggcgcct gaagcgggga
     1741 ccagcagcaa tcggcctggg cccggtcatc ggaattgtca tggcagaagc gctgcgaaag
     1801 ctgggagcag acaaggtgcg agtcaaatgg cccaatgacc tgtatctgca ggatagaaag
     1861 ctggcaggca tcctggtgga gctggccgga ataacaggcg atgctgcaca gatcgtcatt
     1921 ggcgccggga ttaacgtggc tatgaggcgc gtggaggaaa gcgtggtcaa tcagggctgg
     1981 atcacactgc aggaagcagg gattaacctg gacaggaata ctctggccgc tacgctgatc
     2041 cgagagctgc gggcagccct ggaactgttc gagcaggaag gcctggctcc atatctgcca
     2101 cggtgggaga agctggataa cttcatcaat agacccgtga agctgatcat tggggacaaa
     2161 gagattttcg ggattagccg ggggattgat aaacagggag ccctgctgct ggaacaggac
     2221 ggagttatca aaccctggat gggcggagaa atcagtctgc ggtctgccga aaagggctcg
     2281 ggcgggagta ccggtggtag tgggagcggt cgtcgtaagt ggaataagac aggtcacgca
     2341 gtcagagcta taggtcggct gagctcaggt ggaagtggat caggcagcgg tggatctatg
     2401 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     2461 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
     2521 aagctgaccc tgaagttcat ctgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     2581 gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag
     2641 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
     2701 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     2761 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     2821 ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc
     2881 atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac
     2941 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     3001 ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     3061 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtagccc
     3121 ccccccctaa cgttactggc cgaagccgct tggaataagg ccggtgtgcg tttgtctata
     3181 tgttattttc caccatattg ccgtcttttg gcaatgtgag ggcccggaaa cctggccctg
     3241 tcttcttgac gagcattcct aggggtcttt cccctctcgc caaaggaatg caaggtctgt
     3301 tgaatgtcgt gaaggaagca gttcctctgg aagcttcttg aagacaaaca acgtctgtag
     3361 cgaccctttg caggcagcgg aaccccccac ctggcgacag gtgcctctgc ggccaaaagc
     3421 cacgtgtata agatacacct gcaaaggcgg cacaacccca gtgccacgtt gtgagttgga
     3481 tagttgtgga aagagtcaaa tggctctcct caagcgtatt caacaagggg ctgaaggatg
     3541 cccagaaggt accccattgt atgggatctg atctggggcc tcggtgcaca tgctttacat
     3601 gtgtttagtc gaggttaaaa aaacgtctag gccccccgaa ccacggggac gtggttttcc
     3661 tttgaaaaac acgatgataa tatggccaca gccaccatgg accaactgac tgaagagcag
     3721 atcgcagaat ttaaagaggc tttctcccta tttgacaagg acggggatgg gacaataaca
     3781 accaaggagc tggggacggt gatgcggtct ctggggcaga accccacaga agcagagctg
     3841 caggacatga tcaatgaagt agatgccgac ggtgacggca caatcgactt ccctgagttc
     3901 ctgacaatga tggcaagaaa aatgaaatac agggacacgg aagaagaaat tagagaagcg
     3961 ttcggtgtgt ttgataagga tggcaatggc tacatcagtg cagcagagct tcgccacgtg
     4021 atgacaaacc ttggagagaa gttaacagat gaagaggttg atgaaatgat cagggaagca
     4081 gacatcgatg gggatggtca ggtaaactac gaagagtttg tacaaatgat gacagcgaag
     4141 aagggctcgg gctcgacctc gggctcgggc agcggtggct ctggaggttc tggtggcagc
     4201 tctggaggtg gcaagcccat ccccaacccc ctgctgggcc tggacagcac cgctagcaaa
     4261 gacaatactg tgcctctgaa gctgatcgct ctcctggcta atggcgagtt ccatagtggc
     4321 gaacagctgg gagaaaccct gggcatgtcc agggccgcta tcaacaagca cattcagact
     4381 ctgcgcgact ggggcgtgga cgtgttcacc gtgcccggaa agggctactc tctgcccgag
     4441 cctatcccgc tgctgaacgc taaacagatt ctgtaataag aattcctcga gctagagctc
     4501 gctgatcagc ctcgactgtg ccttctagtt gccagccatc tgttgtttgc ccctcccccg
     4561 tgccttcctt gaccctggaa ggtgccactc ccactgtcct ttcctaataa aatgaggaaa
     4621 ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg gggtggggtg gggcaggaca
     4681 gcaaggggga ggattgggaa gacaatagca ggcatgctgg ggaggtaacc acgtgcggac
     4741 cgagcggccg caggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc
     4801 gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc
     4861 agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt ttctccttac
     4921 gcatctgtgc ggtatttcac accgcatacg tcaaagcaac catagtacgc gccctgtagc
     4981 ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc
     5041 gccttagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt
     5101 ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac
     5161 ctcgacccca aaaaacttga tttgggtgat ggttcacgta gtgggccatc gccctgatag
     5221 acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa
     5281 actggaacaa cactcaactc tatctcgggc tattcttttg atttataagg gattttgccg
     5341 atttcggtct attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac
     5401 aaaatattaa cgtttacaat tttatggtgc actctcagta caatctgctc tgatgccgca
     5461 tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg
     5521 ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg
     5581 ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta
     5641 taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat
     5701 gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg
     5761 agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa
     5821 catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac
     5881 ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac
     5941 atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt
     6001 ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc
     6061 gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca
     6121 ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc
     6181 ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag
     6241 gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa
     6301 ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg
     6361 gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa
     6421 ttaatagact ggatggaggt ggataaagtt gcaggaccac ttctgcgctc ggcccttccg
     6481 gctggctggt ttattgctga taaatctgga gccggtgagc gtggaagccg cggtatcatt
     6541 gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt
     6601 caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag
     6661 cattggtaac tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat
     6721 ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct
     6781 taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct
     6841 tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca
     6901 gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc
     6961 agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc
     7021 aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct
     7081 gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag
     7141 gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc
     7201 tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg
     7261 agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag
     7321 cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt
     7381 gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac
     7441 gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgt
//
