LOCUS       sequence-460492-        5819 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Express GFP tagged upsteram ORF of NOTCH2NLC with an expansion 
            (100x) of GGC repeats with codon optimization (NOT pure GGC 
            repeats).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5819)
  AUTHORS   Boivin M, Deng J, Pfister V, Grandgirard E, Oulad-Abdelghani M, 
            Morlet B, Ruffenach F, Negroni L, Koebel P, Jacob H, Riet F, 
            Dijkstra AA, McFadden K, Clayton WA, Hong D, Miyahara H, Iwasaki Y, 
            Sone J, Wang Z, Charlet-Berguerand N
  TITLE     Translation of GGC repeat expansions into a toxic polyglycine 
            protein in NIID defines a novel class of human genetic disorders: 
            The polyG diseases.
  JOURNAL   Neuron. 2021 Jun 2;109(11):1825-1835.e5. doi: 
            10.1016/j.neuron.2021.03.038. Epub 2021 Apr 21.
  PUBMED    33887199
REFERENCE   2  (bases 1 to 5819)
  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: 
            "10.1016/j.neuron.2021.03.038"; journalName: "Neuron"; date: 
            "2021-06-2- 2"; volume: "109"; issue: "11"; pages: "1825-1835.e5"
COMMENT     Sequence Label: pAAV uN2CpolyGly op GFP (NIID)
FEATURES             Location/Qualifiers
     source          1..5819
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     enhancer        206..509
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        510..713
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     664..684
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     710..734
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     intron          862..1240
                     /label=chimeric intron
                     /note="fusion between human cytomegalovirus intron A and 
                     human beta-globin intron 2"
     CDS             join(1766..1768,1769..2482)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(1766..1768,1769..2479)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2069..2088)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2416..2437
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             2525..2551
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     polyA_signal    2566..3042
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2765..2784)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3082..3222
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3082..3211
                     /label=AAV2 ITR
     rep_origin      3297..3752
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3384..3403)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3594..3615
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3769..3788)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3888..3910
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3948..3966)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4034..4138
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4139..4207,4208..4999)
                     /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(4357..4376)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5170..5758
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5659..5678
                     /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 gcgtctagtt attaatagta atcaattacg gggtcattag
      181 ttcatagccc atatatggag ttccgcgtta cataacttac ggtaaatggc ccgcctggct
      241 gaccgcccaa cgacccccgc ccattgacgt caataatgac gtatgttccc atagtaacgc
      301 caatagggac tttccattga cgtcaatggg tggagtattt acggtaaact gcccacttgg
      361 cagtacatca agtgtatcat atgccaagta cgccccctat tgacgtcaat gacggtaaat
      421 ggcccgcctg gcattatgcc cagtacatga ccttatggga ctttcctact tggcagtaca
      481 tctacgtatt agtcatcgct attaccatgg tgatgcggtt ttggcagtac atcaatgggc
      541 gtggatagcg gtttgactca cggggatttc caagtctcca ccccattgac gtcaatggga
      601 gtttgttttg gcaccaaaat caacgggact ttccaaaatg tcgtaacaac tccgccccat
      661 tgacgcaaat gggcggtagg cgtgtacggt gggaggtcta tataagcaga gctcgtttag
      721 tgaaccgtca gatcgcctgg agacgccatc cacgctgttt tgacctccat agaagacacc
      781 gggaccgatc cagcctccgc ggattcgaat cccggccggg aacggtgcat tggaacgcgg
      841 attccccgtg ccaagagtga cgtaagtacc gcctatagag tctataggcc cacaaaaaat
      901 gctttcttct tttaatatac ttttttgttt atcttatttc taatactttc cctaatctct
      961 ttctttcagg gcaataatga tacaatgtat catgcctctt tgcaccattc taaagaataa
     1021 cagtgataat ttctgggtta aggcaatagc aatatttctg catataaata tttctgcata
     1081 taaattgtaa ctgatgtaag aggtttcata ttgctaatag cagctacaat ccagctacca
     1141 ttctgctttt attttatggt tgggataagg ctggattatt ctgagtccaa gctaggccct
     1201 tttgctaatc atgttcatac ctcttatctt cctcccacag ctcctgggca acgtgctggt
     1261 ctgtgtgctg gcccatcact ttggcaaaga attgggattc gaacatcgat tgaattcaaa
     1321 gcttgccgcc atgtggattt gcccaggcgg ggggggaggg ggcgggggag gcggaggcgg
     1381 gggaggcggg ggcggcggag ggggaggggg ggggggagga ggcggcgggg gaggaggcgg
     1441 aggggggggc ggcggcggag ggggcggggg cggaggcggc ggagggggcg gaggcggagg
     1501 cgggggagga ggagggggcg gcggaggcgg aggaggcggg ggagggggcg ggggaggcgg
     1561 cggcgggggc ggggggggag ggggaggagg gggaggcgga ggaggagggg gcgggggcgg
     1621 agggggggga ggcgggggcg ggggagacag ggaggatgcc agacccgcac ctctgtgctg
     1681 tggccggtgc tggagaagcg gctgcgctgc tcgccccccc cgaatgcact gttctgtgga
     1741 gatggcaatg aatcctgtcg gatccgtcag caaaggcgag gagcttttca ccggggttgt
     1801 tcccatcctc gtcgagctcg acggcgacgt aaacggccac aaattcagcg tttccggcga
     1861 gggcgagggc gatgccacct acggcaagct caccctcaag ttcatctgca ccaccggcaa
     1921 gctccccgtc ccctggccca ccctcgtcac caccctcacc tacggcgtcc agtgcttcag
     1981 ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc ccgaaggcta
     2041 cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc gcgccgaggt
     2101 gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg acttcaagga
     2161 ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca acgtctatat
     2221 catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc acaacatcga
     2281 ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg gcgacggccc
     2341 cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca aagaccccaa
     2401 cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga tcactctcgg
     2461 catggacgag ctgtacaagt gataagaatt ctgcagatat ccagcacagt ggcggccgct
     2521 cgagtaccca tacgatgttc cagattacgc taagtaaaga tctacgggtg gcatccctgt
     2581 gacccctccc cagtgcctct cctggccctg gaagttgcca ctccagtgcc caccagcctt
     2641 gtcctaataa aattaagttg catcattttg tctgactagg tgtccttcta taatattatg
     2701 gggtggaggg gggtggtatg gagcaagggg caagttggga agacaacctg tagggcctgc
     2761 ggggtctatt gggaaccaag ctggagtgca gtggcacaat cttggctcac tgcaatctcc
     2821 gcctcctggg ttcaagcgat tctcctgcct cagcctcccg agttgttggg attccaggca
     2881 tgcatgacca ggctcagcta atttttgttt ttttggtaga gacggggttt caccatattg
     2941 gccaggctgg tctccaactc ctaatctcag gtgatctacc caccttggcc tcccaaattg
     3001 ctgggattac aggcgtgaac cactgctccc ttccctgtcc ttctgatttt gtaggtaacc
     3061 acgtgcggac cgagcggccg caggaacccc tagtgatgga gttggccact ccctctctgc
     3121 gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc
     3181 gggcggcctc agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt
     3241 ttctccttac gcatctgtgc ggtatttcac accgcatacg tcaaagcaac catagtacgc
     3301 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     3361 acttgccagc gccttagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     3421 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     3481 tttacggcac ctcgacccca aaaaacttga tttgggtgat ggttcacgta gtgggccatc
     3541 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     3601 cttgttccaa actggaacaa cactcaactc tatctcgggc tattcttttg atttataagg
     3661 gattttgccg atttcggtct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     3721 gaattttaac aaaatattaa cgtttacaat tttatggtgc actctcagta caatctgctc
     3781 tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg cgccctgacg
     3841 ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg ggagctgcat
     3901 gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg
     3961 cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag gtggcacttt
     4021 tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt caaatatgta
     4081 tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa ggaagagtat
     4141 gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt gccttcctgt
     4201 ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg
     4261 agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt ttcgccccga
     4321 agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg tattatcccg
     4381 tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga atgacttggt
     4441 tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa gagaattatg
     4501 cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga caacgatcgg
     4561 aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa ctcgccttga
     4621 tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca ccacgatgcc
     4681 tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta ctctagcttc
     4741 ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac ttctgcgctc
     4801 ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc gtgggtctcg
     4861 cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag ttatctacac
     4921 gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga taggtgcctc
     4981 actgattaag cattggtaac tgtcagacca agtttactca tatatacttt agattgattt
     5041 aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata atctcatgac
     5101 caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa
     5161 aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc
     5221 accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt ttccgaaggt
     5281 aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc cgtagttagg
     5341 ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa tcctgttacc
     5401 agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa gacgatagtt
     5461 accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc ccagcttgga
     5521 gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa gcgccacgct
     5581 tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa caggagagcg
     5641 cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca
     5701 cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa
     5761 cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg ctcacatgt
//
