LOCUS       sequence-350565-        6820 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Viral vector for co-expression of TeNT and EGFP in cells expressing 
            Cre AND NOT Flp driven by the human Synapsin promoter..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6820)
  AUTHORS   Ubina T, Vahedi-Hunter T, Agnew-Svoboda W, Wong W, Gupta A, 
            Santhakumar V, Riccomagno MM
  TITLE     ExBoX - a simple Boolean exclusion strategy to drive expression in 
            neurons.
  JOURNAL   J Cell Sci. 2021 Oct 15;134(20). pii: 272538. doi: 
            10.1242/jcs.257212. Epub 2021 Oct 20.
  PUBMED    34515305
REFERENCE   2  (bases 1 to 6820)
  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: "J Cell 
            Sci."; date: "2021-10-15"; volume: "134(20). pii: 272538. doi"; 
            pages: " 10.1242/jcs.257212. Epub 2021 Oct 20"
COMMENT     Sequence Label: pAAV-SynI-CreOn-FlpOff-TeNT-P2A-EGFP
FEATURES             Location/Qualifiers
     source          1..6820
                     /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"
     protein_bind    complement(646..679)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(733..766)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    773..806
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     CDS             complement(join(850..1563,1564..1566,1567..1569))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGDELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(895..916)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1244..1263
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1503..1524
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             complement(1570..1626)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     protein_bind    3049..3082
                     /label=FRT (minimal)
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision but not integration 
                     (Turan and Bode, 2011)"
     protein_bind    3092..3125
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    3176..3209
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(3210..3234)
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     misc_feature    3244..3832
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3297..3317)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3715..3726)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(3862..3879)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    3868..4075
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   4083..4223
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4083..4212
                     /label=AAV2 ITR
     rep_origin      4298..4753
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4385..4404)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4595..4616
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4770..4789)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4889..4911
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4949..4967)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5035..5139
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5140..5208,5209..6000)
                     /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(5358..5377)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6171..6759
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6660..6679
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcggc
       61 ctcagtgagc gagcgagcgc gcagagaggg agtggccaac tccatcacta ggggttcctt
      121 ctagacaact 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 ccaccataac ttcgtatagg
      661 atactttata cgaagttatc cggcagaatg gtagctggat tgtagctgct attagcaata
      721 tgaaacctct taataacttc gtatagcata cattatacga agttatggcg cggaagttcc
      781 tattctctag aaagtatagg aacttccctc gagtaccggt cacgcgtacg caagcgcctt
      841 agcggccgct tacttgtaca gctcgtccat gccgagagtg atcccggcgg cggtcacgaa
      901 ctccagcagg accatgtgat cgcgcttctc gttggggtct ttgctcaggg cggactgggt
      961 gctcaggtag tggttgtcgg gcagcagcac ggggccgtcg ccgatggggg tgttctgctg
     1021 gtagtggtcg gcgagctgca cgctgccgtc ctcgatgttg tggcggatct tgaagttcac
     1081 cttgatgccg ttcttctgct tgtcggccat gatatagacg ttgtggctgt tgtagttgta
     1141 ctccagcttg tgccccagga tgttgccgtc ctccttgaag tcgatgccct tcagctcgat
     1201 gcggttcacc agggtgtcgc cctcgaactt cacctcggcg cgggtcttgt agttgccgtc
     1261 gtccttgaag aagatggtgc gctcctggac gtagccttcg ggcatggcgg acttgaagaa
     1321 gtcgtgctgc ttcatgtggt cggggtagcg gctgaagcac tgcacgccgt aggtcagggt
     1381 ggtcacgagg gtgggccagg gcacgggcag cttgccggtg gtgcagatga acttcagggt
     1441 cagcttgccg taggtggcat cgccctcgcc ctcgccggac acgctgaact tgtggccgtt
     1501 tacgtcgccg tccagctcga ccaggatggg caccaccccg gtgaacagct cgtcgccctt
     1561 gctcaccata ggcccggggt tttcttcaac atctcctgct tgctttaaca gagagaagtt
     1621 cgtggctccg cttccagcgg tacggttgta caggttttca cggatgttgg tcggcgggat
     1681 gatttttttg cacagaccga tcagtttgga aaccagaccg gaaccgtcga cgttcctaaa
     1741 ggcgttagtg ttcaccctca tgttctggcc cttatactcg gacttcagat ccttggactc
     1801 tatgttaaag ccctcagtat cgttatatat agtatcatcc agcaggttag gtatcttcac
     1861 aggatcgtgg ttcatggaaa aatagcttaa gcgagtcttt atgttaaact tcttgcccag
     1921 ctctatctca gtaaagccat acattatgga gttatacagt atctgaaact tatcctcgtt
     1981 cactatatac tggccgttgg aatccttatc aaactgatac ttctgctgat agatctgttt
     2041 gtaggagtcg atgtcgatgt tcgggtcgtt gcaggaggta acctgggaca gtttgttagc
     2101 gatagctttg tagtcgttca gggttttttc gtacaggtcg tttttgatat cgatggagat
     2161 caggttagcg tcctgaccac cgaaggtgaa gagctcttca gcggagatcg ggtaggtgtg
     2221 ctgcatgtag atttcctgtt tggacgggat gatttcgtgg gaggaaacct gcatgccgta
     2281 cagaccgtgc agaacgtgga tcagttcgtg catcagcagc agagccgggt cctggaagta
     2341 tttggattta ccgatggtca gggaggtgat gttttcgata acgttgtcga aggtcggtac
     2401 gtattccggg cagaaagcca tctgcatgat ggaaccgaaa ccgtcacggc acgggaagta
     2461 gtttttgttg tcaacacgca gaacgatacc acgaacttcg tttttgttca gaaccggacc
     2521 cggaccgaag atgatcaggt tggtcagcat agcggatttg gtggtagcac cggacggatc
     2581 ctgttccagc aggttgaagg aaacggagtt ggagttggtg tcgaatttgt ccagcaggga
     2641 gtaggagtta cccaggtacg ggatagcgtt gatgattttg tccagcagag cttcaccagc
     2701 aacgttgttt ttgatacggt tgaacagttt aaccatggtc tgcaggaaac ggtctttgtc
     2761 ggagtcggta cgcaggtagt tcgggtcgta gtattcggaa gcaccttcga tcagggagga
     2821 cggcgggttg aagtcttccg gtttggtacc aaattcgtaa cgttccggaa cgatccagat
     2881 acggtcggtg attttgaagg ctttgtagta gatgtctaga cctttgcagt acggcggttc
     2941 catcatgatg atggtgtcgt tgttaaccgg gtcggagtaa cggaagttgt tgatggtgat
     3001 cggcatggtg gcggatccta ggtacctgag ctcaacgata gcactagtga agttcctatt
     3061 ctctagaaag tataggaact tcgctagccg gataacttcg tataaagtat cctatacgaa
     3121 gttatttgcc ttaacccaga aattatcact gttattcttt agaatggtgc aaagaataac
     3181 ttcgtataat gtatgctata cgaagttata cccagctttc ttgtacaaag tgggaattcc
     3241 gataatcaac ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt
     3301 gctcctttta cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc
     3361 cgtatggctt tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag
     3421 ttgtggcccg ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc
     3481 actggttggg gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc
     3541 cctattgcca cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg
     3601 ctgttgggca ctgacaattc cgtggtgttg tcggggaagc tgacgtcctt tccatggctg
     3661 ctcgcctgtg ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc
     3721 ctcaatccag cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt
     3781 cttcgccttc gccctcagac gagtcggatc tccctttggg ccgcctcccc gcatcgggaa
     3841 ttcctagagc tcgctgatca gcctcgactg tgccttctag ttgccagcca tctgttgttt
     3901 gcccctcccc cgtgccttcc ttgaccctgg aaggtgccac tcccactgtc ctttcctaat
     3961 aaaatgagga aattgcatcg cattgtctga gtaggtgtca ttctattctg gggggtgggg
     4021 tggggcagga cagcaagggg gaggattggg aagagaatag caggcatgct ggggagggcc
     4081 gcaggaaccc ctagtgatgg agttggccac tccctctctg cgcgctcgct cgctcactga
     4141 ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc cgggcggcct cagtgagcga
     4201 gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat tttctcctta cgcatctgtg
     4261 cggtatttca caccgcatac gtcaaagcaa ccatagtacg cgccctgtag cggcgcatta
     4321 agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta cacttgccag cgccttagcg
     4381 cccgctcctt tcgctttctt cccttccttt ctcgccacgt tcgccggctt tccccgtcaa
     4441 gctctaaatc gggggctccc tttagggttc cgatttagtg ctttacggca cctcgacccc
     4501 aaaaaacttg atttgggtga tggttcacgt agtgggccat cgccctgata gacggttttt
     4561 cgccctttga cgttggagtc cacgttcttt aatagtggac tcttgttcca aactggaaca
     4621 acactcaact ctatctcggg ctattctttt gatttataag ggattttgcc gatttcggtc
     4681 tattggttaa aaaatgagct gatttaacaa aaatttaacg cgaattttaa caaaatatta
     4741 acgtttacaa ttttatggtg cactctcagt acaatctgct ctgatgccgc atagttaagc
     4801 cagccccgac acccgccaac acccgctgac gcgccctgac gggcttgtct gctcccggca
     4861 tccgcttaca gacaagctgt gaccgtctcc gggagctgca tgtgtcagag gttttcaccg
     4921 tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac gcctattttt ataggttaat
     4981 gtcatgataa taatggtttc ttagacgtca ggtggcactt ttcggggaaa tgtgcgcgga
     5041 acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat gagacaataa
     5101 ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca acatttccgt
     5161 gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca cccagaaacg
     5221 ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta catcgaactg
     5281 gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt tccaatgatg
     5341 agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc cgggcaagag
     5401 caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc accagtcaca
     5461 gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc cataaccatg
     5521 agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa ggagctaacc
     5581 gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga accggagctg
     5641 aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat ggcaacaacg
     5701 ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca attaatagac
     5761 tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc ggctggctgg
     5821 tttattgctg ataaatctgg agccggtgag cgtggaagcc gcggtatcat tgcagcactg
     5881 gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag tcaggcaact
     5941 atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa gcattggtaa
     6001 ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca tttttaattt
     6061 aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc ttaacgtgag
     6121 ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc ttgagatcct
     6181 ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc agcggtggtt
     6241 tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt cagcagagcg
     6301 cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt caagaactct
     6361 gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc tgccagtggc
     6421 gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa ggcgcagcgg
     6481 tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac ctacaccgaa
     6541 ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg gagaaaggcg
     6601 gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga gcttccaggg
     6661 ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact tgagcgtcga
     6721 tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa cgcggccttt
     6781 ttacggttcc tggccttttg ctggcctttt gctcacatgt
//
