LOCUS       sequence-370111-        5744 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes a specific PSD-95 binder (Xph20) fused to eGFP, CCR5 
            left-handed zinc finger, and KRAB(A) transcriptional repressor 
            domain.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5744)
  AUTHORS   Rimbault C, Breillat C, Compans B, Toulme E, Nunes Vicente F, 
            Fernandez-Monreal M, Mascalchi P, Genuer C, Puente-Munoz V, 
            Gauthereau I, Hosy E, Claverol S, Giannone G, Chamma I, Mackereth 
            CD, Poujol C, Choquet D, Sainlos M
  TITLE     Engineering paralog-specific PSD-95 recombinant binders as minimally
            interfering multimodal probes for advanced imaging techniques.
  JOURNAL   Elife. 2024 Jan 3;13:e69620. doi: 10.7554/eLife.69620.
  PUBMED    38167295
REFERENCE   2  (bases 1 to 5744)
  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: "Elife."; 
            date: "2024-01-3"; volume: "13:e69620. doi"; pages: " 
            10.7554/eLife.69620"
COMMENT     Sequence Label: AAV_Syn_Xph20_eGFP_CCR5TC
FEATURES             Location/Qualifiers
     source          1..5744
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     protein_bind    196..207
                     /label=ZF binding site
                     /bound_moiety="CCR5TC zinc-finger domain"
                     /note="target sequence for the CCR5TC zinc-finger domain 
                     from the CCR5-224 (+) zinc-finger nuclease (Perez et al., 
                     2008; Gross et al., 2013)"
     promoter        235..682
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     regulatory      705..714
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(1029..1031,1032..1739)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELY"
     CDS             join(1029..1031,1032..1739)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELY"
     primer_bind     complement(1071..1092)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1332..1351)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1679..1700
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1785..2126
                     /codon_start=1
                     /product="zinc-finger domain from the CCR5-224 (+) 
                     zinc-finger nuclease that recognizes the left half-site 
                     target sequence in the human CCR5 gene (Perez et al., 2008;
                     Gross et al., 2013)"
                     /label=CCR5TC
                     /translation="FQCRICMRNFSDRSNLSRHIRTHTGEKPFACDICGRKFAISSNLN
                     SHTKIHTGSQKPFQCRICMRNFSRSDNLARHIRTHTGEKPFACDICGRKFATSGNLTRH
                     TKIHLRGSQL"
     CDS             2175..2300
                     /codon_start=1
                     /product="Kruppel-associated box (KRAB) transcriptional 
                     repression domain from the rat zinc finger protein Kid-1 
                     (Witzgall et al., 1994)"
                     /label=KRAB-A
                     /translation="VSVTFEDVAVLFTRDEWKKLDLSQRSLYREVMLENYSNLASM"
     misc_feature    2331..2919
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2384..2404)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2802..2813)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2922..2938)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2923..2939)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    2968..3089
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2979..2998)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3033..3052
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(3468..4056)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3548..3567)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4227..5018,5019..5087))
                     /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     4850..4869
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5088..5192)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5218..5673)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5355..5376)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5567..5586
                     /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 aggaagatca cgcgtgtcat cctcatcaga tctctgcaga gggccctgcg tatgagtgca
      241 agtgggtttt aggaccagga tgaggcgggg tgggggtgcc tacctgacga ccgaccccga
      301 cccactggac aagcacccaa cccccattcc ccaaattgcg catcccctat cagagagggg
      361 gaggggaaac aggatgcggc gaggcgcgtg cgcactgcca gcttcagcac cgcggacagt
      421 gccttcgccc ccgcctggcg gcgcgcgcca ccgccgcctc agcactgaag gcgcgctgac
      481 gtcactcgcc ggtcccccgc aaactcccct tcccggccac cttggtcgcg tccgcgccgc
      541 cgccggccca gccggaccgc accacgcgag gcgcgagata ggggggcacg ggcgcgacca
      601 tctgcgctgc ggcgccggcg actcagcgct gcctcagtct gcggtgggca gcggaggagt
      661 cgtgtcgtgc ctgagagcgc agtcgaattc aagctgctag cgccgccacc atggcgggca
      721 gctctgtcag ttccgtgccg accaaactgg aagtggttgc ggccaccccg acgtctctgc
      781 tgatcagctg ggatgcagtg gcgaagaatg tgaaggtcgg ttattaccgt atcacctatg
      841 gcgaaacggg cggtaactcc ccggttcagg aattcaccgt cccgggtagt tcctcaaccg
      901 caacgatttc cggcctgtca ccgggtgtcg actataccat tacggtttac gctaatgggg
      961 tgctgagtaa gatggtgctt ccgatttcta tcaattaccg taccggtagc ggctccagta
     1021 gatctggggt gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc
     1081 tggacggcga cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca
     1141 cctacggcaa gctgaccctg aagttcatct gcaccaccgg caagctgccc gtgccctggc
     1201 ccaccctcgt gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca
     1261 tgaagcagca cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca
     1321 tcttcttcaa ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca
     1381 ccctggtgaa ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg
     1441 ggcacaagct ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga
     1501 agaacggcat caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc
     1561 tcgccgacca ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca
     1621 accactacct gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca
     1681 tggtcctgct ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtact
     1741 ctaggggagc tggcgctgga gcgggtgcag gggctggctc tagattccag tgccggatct
     1801 gcatgcggaa cttcagcgac cggtccaacc tgagcaggca catcagaacc cacaccggag
     1861 aaaagccctt cgcctgcgac atttgcggcc ggaagttcgc catcagcagc aacctgaaca
     1921 gccacaccaa gatccacact ggcagccaga aacctttcca gtgcagaatt tgtatgagaa
     1981 actttagcag aagcgacaac ctggccagac acatccggac acatactggt gaaaaacctt
     2041 ttgcctgtga tatctgtggc agaaagtttg ccacctccgg caatctgacc cggcacacaa
     2101 agattcacct gcggggcagc cagctatcga ttgtcgacgc tcctgaacaa cgtgaaggtg
     2161 cttctcaagt ttctgtttct gttacttttg aagatgttgc tgttcttttt actcgtgatg
     2221 aatggaaaaa acttgatctt tctcaacgtt ctctttatcg tgaagttatg cttgaaaatt
     2281 attctaatct tgcttctatg gcttaagaat tcgatatcaa gcttatcgat aatcaacctc
     2341 tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc
     2401 tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca
     2461 ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg
     2521 tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca
     2581 ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg
     2641 cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg
     2701 acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctgtgttg
     2761 ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg
     2821 accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc
     2881 ctcagacgag tcggatctcc ctttgggccg cctccccgca tcgataccgt cgacctcgac
     2941 ccgggcggcc gcttcgagca gacatgataa gatacattga tgagtttgga caaaccacaa
     3001 ctagaatgca gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt gctttatttg
     3061 taaccattat aagctgcaat aaacaagtta acaacaacaa ttgcattcat tttatgtttc
     3121 aggttcaggg ggagatgtgg gaggtttttt aaagcaagta aaacctctac aaatgtggta
     3181 aaatcgataa ggatcttcct agagcatggc tacgtagata agtagcatgg cgggttaatc
     3241 attaactaca aggaacccct agtgatggag ttggccactc cctctctgcg cgctcgctcg
     3301 ctcactgact cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag
     3361 gcggtaatac ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa
     3421 ggccagcaaa aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc
     3481 cgcccccctg acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca
     3541 ggactataaa gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg
     3601 accctgccgc ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct
     3661 catagctcac gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt
     3721 gtgcacgaac cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag
     3781 tccaacccgg taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc
     3841 agagcgaggt atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac
     3901 actagaagaa cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga
     3961 gttggtagct cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc
     4021 aagcagcaga ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg
     4081 gggtctgacg ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca
     4141 aaaaggatct tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt
     4201 atatatgagt aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca
     4261 gcgatctgtc tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg
     4321 atacgggagg gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca
     4381 ccggctccag atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt
     4441 cctgcaactt tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt
     4501 agttcgccag ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca
     4561 cgctcgtcgt ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca
     4621 tgatccccca tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga
     4681 agtaagttgg ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact
     4741 gtcatgccat ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga
     4801 gaatagtgta tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg
     4861 ccacatagca gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc
     4921 tcaaggatct taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga
     4981 tcttcagcat cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat
     5041 gccgcaaaaa agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt
     5101 caatattatt gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt
     5161 atttagaaaa ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctaaa
     5221 ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc agctcatttt
     5281 ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag accgagatag
     5341 ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg gactccaacg
     5401 tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca tcaccctaat
     5461 caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa gggagccccc
     5521 gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg aagaaagcga
     5581 aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta accaccacac
     5641 ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gccattcagg ctgcgcaact
     5701 gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccag
//
