LOCUS       sequence-370109-        5747 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes a specific PSD-95 binder (Xph18) 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 5747)
  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 5747)
  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_Xph18_eGFP_CCR5TC
FEATURES             Location/Qualifiers
     source          1..5747
                     /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(1032..1034,1035..1742)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELY"
     CDS             join(1032..1034,1035..1742)
                     /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(1074..1095)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1335..1354)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1682..1703
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1788..2129
                     /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             2178..2303
                     /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    2334..2922
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2387..2407)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2805..2816)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2925..2941)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2926..2942)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    2971..3092
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2982..3001)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     3036..3055
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     rep_origin      complement(3471..4059)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3551..3570)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(4230..5021,5022..5090))
                     /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     4853..4872
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(5091..5195)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(5221..5676)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5358..5379)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5570..5589
                     /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 ggatgcacat tcgcggatgg cgtgggtcaa ttattaccgt atcacctatg
      841 gcgaaacggg cggtaactcc ccggttcagg aattcaccgt cccgggtagt tcctcaaccg
      901 caacgatttc cggcctgtca ccgggtgtcg actataccat tacggtttac gctaatgggg
      961 gggggtttcg tcgggggatg gcgccgattt ctatcaatta ccgtaccggt agcggctcca
     1021 gtagatctgg ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg
     1081 agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg
     1141 ccacctacgg caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct
     1201 ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc
     1261 acatgaagca gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca
     1321 ccatcttctt caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg
     1381 acaccctggt gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc
     1441 tggggcacaa gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc
     1501 agaagaacgg catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc
     1561 agctcgccga ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg
     1621 acaaccacta cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc
     1681 acatggtcct gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt
     1741 actctagggg agctggcgct ggagcgggtg caggggctgg ctctagattc cagtgccgga
     1801 tctgcatgcg gaacttcagc gaccggtcca acctgagcag gcacatcaga acccacaccg
     1861 gagaaaagcc cttcgcctgc gacatttgcg gccggaagtt cgccatcagc agcaacctga
     1921 acagccacac caagatccac actggcagcc agaaaccttt ccagtgcaga atttgtatga
     1981 gaaactttag cagaagcgac aacctggcca gacacatccg gacacatact ggtgaaaaac
     2041 cttttgcctg tgatatctgt ggcagaaagt ttgccacctc cggcaatctg acccggcaca
     2101 caaagattca cctgcggggc agccagctat cgattgtcga cgctcctgaa caacgtgaag
     2161 gtgcttctca agtttctgtt tctgttactt ttgaagatgt tgctgttctt tttactcgtg
     2221 atgaatggaa aaaacttgat ctttctcaac gttctcttta tcgtgaagtt atgcttgaaa
     2281 attattctaa tcttgcttct atggcttaag aattcgatat caagcttatc gataatcaac
     2341 ctctggatta caaaatttgt gaaagattga ctggtattct taactatgtt gctcctttta
     2401 cgctatgtgg atacgctgct ttaatgcctt tgtatcatgc tattgcttcc cgtatggctt
     2461 tcattttctc ctccttgtat aaatcctggt tgctgtctct ttatgaggag ttgtggcccg
     2521 ttgtcaggca acgtggcgtg gtgtgcactg tgtttgctga cgcaaccccc actggttggg
     2581 gcattgccac cacctgtcag ctcctttccg ggactttcgc tttccccctc cctattgcca
     2641 cggcggaact catcgccgcc tgccttgccc gctgctggac aggggctcgg ctgttgggca
     2701 ctgacaattc cgtggtgttg tcggggaaat catcgtcctt tccttggctg ctcgcctgtg
     2761 ttgccacctg gattctgcgc gggacgtcct tctgctacgt cccttcggcc ctcaatccag
     2821 cggaccttcc ttcccgcggc ctgctgccgg ctctgcggcc tcttccgcgt cttcgccttc
     2881 gccctcagac gagtcggatc tccctttggg ccgcctcccc gcatcgatac cgtcgacctc
     2941 gacccgggcg gccgcttcga gcagacatga taagatacat tgatgagttt ggacaaacca
     3001 caactagaat gcagtgaaaa aaatgcttta tttgtgaaat ttgtgatgct attgctttat
     3061 ttgtaaccat tataagctgc aataaacaag ttaacaacaa caattgcatt cattttatgt
     3121 ttcaggttca gggggagatg tgggaggttt tttaaagcaa gtaaaacctc tacaaatgtg
     3181 gtaaaatcga taaggatctt cctagagcat ggctacgtag ataagtagca tggcgggtta
     3241 atcattaact acaaggaacc cctagtgatg gagttggcca ctccctctct gcgcgctcgc
     3301 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     3361 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     3421 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     3481 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     3541 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     3601 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     3661 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     3721 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     3781 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     3841 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     3901 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     3961 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     4021 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     4081 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     4141 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     4201 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     4261 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     4321 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     4381 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     4441 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     4501 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     4561 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     4621 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     4681 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     4741 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     4801 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     4861 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4921 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     4981 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     5041 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     5101 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     5161 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     5221 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     5281 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     5341 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
     5401 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
     5461 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
     5521 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
     5581 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
     5641 cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc aggctgcgca
     5701 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccag
//
