LOCUS       sequence-394041-        5641 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses cysteine-free GFP (cfGFP) under synapsin promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5641)
  AUTHORS   Ramsaran AI, Wang Y, Golbabaei A, Aleshin S, de Snoo ML, Yeung BA, 
            Rashid AJ, Awasthi A, Lau J, Tran LM, Ko SY, Abegg A, Duan LC, 
            McKenzie C, Gallucci J, Ahmed M, Kaushik R, Dityatev A, Josselyn SA,
            Frankland PW
  TITLE     A shift in the mechanisms controlling hippocampal engram formation 
            during brain maturation.
  JOURNAL   Science. 2023 May 5;380(6644):543-551. doi: 10.1126/science.ade6530.
            Epub 2023 May 4.
  PUBMED    37141366
REFERENCE   2  (bases 1 to 5641)
  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: 
            "10.1126/science.ade6530"; journalName: "Science"; date: "2023-05-5-
            5"; volume: "380"; issue: "6644"; pages: "543-551"
COMMENT     Sequence Label: AAV-cfGFP
FEATURES             Location/Qualifiers
     source          1..5641
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    126..146
                     /gene="mutant version of attB"
                     /label=attB4
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     promoter        168..615
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    615..639
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     CDS             670..735
                     /codon_start=1
                     /product="three tandem FLAG(R) epitope tags, followed by an
                     enterokinase cleavage site"
                     /label=3xFLAG
                     /translation="DYKDHDGDYKDHDIDYKDDDDK"
     regulatory      757..766
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(763..765,766..768,769..1479)
                     /codon_start=1
                     /product="strongly enhanced green fluorescent protein 
                     (Kremers et al., 2007)"
                     /label=SGFP2
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFISTTGKLPVPWPTLVTTLTYGVQMFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYITADKQKNGIK
                     ANFKIRHNIEDGGVQLADHYQQNTPIGDGPVLLPDNHYLSTQSKLSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(808..829)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1069..1088)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1416..1437
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(1489..1513)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    1517..2105
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1570..1590)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1988..1999)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     protein_bind    complement(2111..2131)
                     /gene="mutant version of attB"
                     /label=attB3
                     /bound_moiety="BP Clonase(TM)"
                     /note="core recombination site for the Gateway(R) BP 
                     reaction"
     polyA_signal    2159..2280
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2170..2189)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2224..2243
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   2384..2524
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2384..2513
                     /label=AAV2 ITR
     primer_bind     complement(2534..2551)
                     /label=M13 Forward
                     /note="In lacZ gene. Also called M13-F20 or M13 (-21) 
                     Forward"
     primer_bind     complement(2534..2550)
                     /label=M13 fwd
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2543..2565)
                     /label=M13/pUC Forward
                     /note="In lacZ gene"
     rep_origin      2763..3218
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2850..2869)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3060..3081
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3235..3254)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3354..3376
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3414..3432)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3500..3604
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3605..3673,3674..4465)
                     /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(3823..3842)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4636..5224
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5125..5144
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     protein_bind    5512..5533
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5548..5553,5554..5571,5572..5578)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5586..5602
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     primer_bind     5591..5613
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5610..5626
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5610..5626
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcggc
       61 ctcagtgagc gagcgagcgc gcagagaggg agtggccaac tccatcacta ggggttccta
      121 tcgatcaact ttgtatagaa aagttgctgc agagggccct gcgtatgagt gcaagtgggt
      181 tttaggacca ggatgaggcg gggtgggggt gcctacctga cgaccgaccc cgacccactg
      241 gacaagcacc caacccccat tccccaaatt gcgcatcccc tatcagagag ggggagggga
      301 aacaggatgc ggcgaggcgc gtgcgcactg ccagcttcag caccgcggac agtgccttcg
      361 cccccgcctg gcggcgcgcg ccaccgccgc ctcagcactg aaggcgcgct gacgtcactc
      421 gccggtcccc cgcaaactcc ccttcccggc caccttggtc gcgtccgcgc cgccgccggc
      481 ccagccggac cgcaccacgc gaggcgcgag ataggggggc acgggcgcga ccatctgcgc
      541 tgcggcgccg gcgactcagc gctgcctcag tctgcggtgg gcagcggagg agtcgtgtcg
      601 tgcctgagag cgcagcaagt ttgtacaaaa aagcaggctg aattcaagct tctgcagctc
      661 gaggctagcg actacaaaga ccatgacggt gattataaag atcatgacat cgactacaag
      721 gatgacgatg acaagtaggg taccgagctc gtcgacgcca ccatggtgag caagggcgag
      781 gagctgttca ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac
      841 aagttcagcg tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag
      901 ttcatctcca ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc
      961 tacggcgtgc agatgttcgc ccgctacccc gaccacatga agcagcacga cttcttcaag
     1021 tccgccatgc ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac
     1081 tacaagaccc gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg
     1141 aagggcatcg acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac
     1201 aacagccaca acgtctatat cacggccgac aagcagaaga acggcatcaa ggccaacttc
     1261 aagatccgcc acaacatcga ggacggcggc gtgcagctcg ccgaccacta ccagcagaac
     1321 acccccatcg gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc
     1381 aaactgagca aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc
     1441 gccgccggga tcactctcgg catggacgag ctgtacaagt aatctagaac ccagctttct
     1501 tgtacaaagt ggcgataatc aacctctgga ttacaaaatt tgtgaaagat tgactggtat
     1561 tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc ctttgtatca
     1621 tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct ggttgctgtc
     1681 tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca ctgtgtttgc
     1741 tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt ccgggacttt
     1801 cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg cccgctgctg
     1861 gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga agctgacgtc
     1921 ctttccatgg ctgctcgcct gtgttgccac ctggattctg cgcgggacgt ccttctgcta
     1981 cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc cggctctgcg
     2041 gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt gggccgcctc
     2101 cccgcatcgg caactttatt atacatagtt gatggccggc cgcttcgagc agacatgata
     2161 agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa atgctttatt
     2221 tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa taaacaagtt
     2281 aacaacaaca attgcattca ttttatgttt caggttcagg gggaggtgtg ggaggttttt
     2341 taaagcaagt aaaacctcta caaatgtggt aatcgataga tctaggaacc cctagtgatg
     2401 gagttggcca ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc
     2461 gcccgacgcc cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg
     2521 caggcagctt ggcactggcc gtcgttttac aacgtcgtga ctgggaaaac cctggcgtta
     2581 cccaacttaa tcgccttgca gcacatcccc ctttcgccag ctggcgtaat agcgaagagg
     2641 cccgcaccga tcgcccttcc caacagttgc gcagcctgaa tggcgaatgg cgcctgatgc
     2701 ggtattttct ccttacgcat ctgtgcggta tttcacaccg catacgtcaa agcaaccata
     2761 gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg gtggttacgc gcagcgtgac
     2821 cgctacactt gccagcgccc tagcgcccgc tcctttcgct ttcttccctt cctttctcgc
     2881 cacgttcgcc ggctttcccc gtcaagctct aaatcggggg ctccctttag ggttccgatt
     2941 tagtgcttta cggcacctcg accccaaaaa acttgatttg ggtgatggtt cacgtagtgg
     3001 gccatcgccc tgatagacgg tttttcgccc tttgacgttg gagtccacgt tctttaatag
     3061 tggactcttg ttccaaactg gaacaacact caaccctatc tcgggctatt cttttgattt
     3121 ataagggatt ttgccgattt cggcctattg gttaaaaaat gagctgattt aacaaaaatt
     3181 taacgcgaat tttaacaaaa tattaacgtt tacaatttta tggtgcactc tcagtacaat
     3241 ctgctctgat gccgcatagt taagccagcc ccgacacccg ccaacacccg ctgacgcgcc
     3301 ctgacgggct tgtctgctcc cggcatccgc ttacagacaa gctgtgaccg tctccgggag
     3361 ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc gcgagacgaa agggcctcgt
     3421 gatacgccta tttttatagg ttaatgtcat gataataatg gtttcttaga cgtcaggtgg
     3481 cacttttcgg ggaaatgtgc gcggaacccc tatttgttta tttttctaaa tacattcaaa
     3541 tatgtatccg ctcatgagac aataaccctg ataaatgctt caataatatt gaaaaaggaa
     3601 gagtatgagt attcaacatt tccgtgtcgc ccttattccc ttttttgcgg cattttgcct
     3661 tcctgttttt gctcacccag aaacgctggt gaaagtaaaa gatgctgaag atcagttggg
     3721 tgcacgagtg ggttacatcg aactggatct caacagcggt aagatccttg agagttttcg
     3781 ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt ctgctatgtg gcgcggtatt
     3841 atcccgtatt gacgccgggc aagagcaact cggtcgccgc atacactatt ctcagaatga
     3901 cttggttgag tactcaccag tcacagaaaa gcatcttacg gatggcatga cagtaagaga
     3961 attatgcagt gctgccataa ccatgagtga taacactgcg gccaacttac ttctgacaac
     4021 gatcggagga ccgaaggagc taaccgcttt tttgcacaac atgggggatc atgtaactcg
     4081 ccttgatcgt tgggaaccgg agctgaatga agccatacca aacgacgagc gtgacaccac
     4141 gatgcctgta gcaatggcaa caacgttgcg caaactatta actggcgaac tacttactct
     4201 agcttcccgg caacaattaa tagactggat ggaggcggat aaagttgcag gaccacttct
     4261 gcgctcggcc cttccggctg gctggtttat tgctgataaa tctggagccg gtgagcgtgg
     4321 gtctcgcggt atcattgcag cactggggcc agatggtaag ccctcccgta tcgtagttat
     4381 ctacacgacg gggagtcagg caactatgga tgaacgaaat agacagatcg ctgagatagg
     4441 tgcctcactg attaagcatt ggtaactgtc agaccaagtt tactcatata tactttagat
     4501 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     4561 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     4621 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     4681 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     4741 gaaggtaact ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta
     4801 gttaggccac cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct
     4861 gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg
     4921 atagttaccg gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag
     4981 cttggagcga acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc
     5041 cacgcttccc gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg
     5101 agagcgcacg agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt
     5161 tcgccacctc tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg
     5221 gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca
     5281 catgttcttt cctgcgttat cccctgattc tgtggataac cgtattaccg cctttgagtg
     5341 agctgatacc gctcgccgca gccgaacgac cgagcgcagc gagtcagtga gcgaggaagc
     5401 ggaagagcgc ccaatacgca aaccgcctct ccccgcgcgt tggccgattc attaatgcag
     5461 ctggcacgac aggtttcccg actggaaagc gggcagtgag cgcaacgcaa ttaatgtgag
     5521 ttagctcact cattaggcac cccaggcttt acactttatg cttccggctc gtatgttgtg
     5581 tggaattgtg agcggataac aatttcacac aggaaacagc tatgaccatg attacgaatt
     5641 g
//
