LOCUS       sequence-306219-        8628 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  synthetic circular DNA
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 8628)
  AUTHORS   Lopez ME, Klein AD, Dimbil UJ, Scott MP
  TITLE     Anatomically defined neuron-based rescue of neurodegenerative 
            Niemann-Pick type C disorder.
  JOURNAL   J Neurosci. 2011 Mar 23;31(12):4367-78. doi: 
            10.1523/JNEUROSCI.5981-10.2011.
  PUBMED    21430138
REFERENCE   2  (bases 1 to 8628)
  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: 
            "10.1523/JNEUROSCI.5981-10"; journalName: "J Neurosci"; date: 
            "2011-03-23- 23"; volume: "31"; issue: "12"; pages: "4367-78"
COMMENT     Sequence Label: NPC1 His6 EGFP
FEATURES             Location/Qualifiers
     source          1..8628
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     enhancer        1..304
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        305..508
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     459..479
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     CDS             4466..4483
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     regulatory      4508..4517
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(4514..4516,4517..4519,4520..5233)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(4514..4516,4517..4519,4520..5230)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(4559..4580)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(4820..4839)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     5167..5188
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    5356..5477
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5393..5412)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     5447..5466
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(5484..5939)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5621..5642)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     5833..5852
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        5966..6070
                     /gene="bla"
                     /label=AmpR promoter
     promoter        6072..6429
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        6072..6263
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(6095..6115)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      6280..6415
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             6464..7258
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             6464..7258
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(6518..6537)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     7128..7147
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(7446..7465)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    7490..7537
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      7866..8454
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     8355..8374
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cgttacataa cttacggtaa atggcccgcc tggctgaccg cccaacgacc cccgcccatt
       61 gacgtcaata atgacgtatg ttcccatagt aacgccaata gggactttcc attgacgtca
      121 atgggtggag tatttacggt aaactgccca cttggcagta catcaagtgt atcatatgcc
      181 aagtacgccc cctattgacg tcaatgacgg taaatggccc gcctggcatt atgcccagta
      241 catgacctta tgggactttc ctacttggca gtacatctac gtattagtca tcgctattac
      301 catggtgatg cggttttggc agtacatcaa tgggcgtgga tagcggtttg actcacgggg
      361 atttccaagt ctccacccca ttgacgtcaa tgggagtttg ttttggcacc aaaatcaacg
      421 ggactttcca aaatgtcgta acaactccgc cccattgacg caaatgggcg gtaggcgtgt
      481 acggtgggag gtctatataa gcagagctgg tttagtgaac cgtcagatcc gctagcgcta
      541 ccggactcag atctcgagcg gccgccagtg tgatggatat ctgcagaatt cggcttgagc
      601 cgaactgaga gctgtagccc cgcgcggcga cagcatgggt gcgcaccacc cggccctcgg
      661 cctgctgctg ctgctgctgt gccctgcgca ggtgttttcg caatcctgtg tttggtatgg
      721 agagtgtgga attgcgactg gagataagag gtacaactgt aaatattctg gcccaccaaa
      781 acccctccca aaggacggct atgacttagt gcaggaactc tgtccaggac tcttctttga
      841 caatgtcagt ctctgctgtg acattcaaca gcttcagacg ctgaagagta acctgcagct
      901 gcccctgcag ttcctgtcca ggtgtccgtc atgtttttat aacctaatga ccctgttttg
      961 tgagctaaca tgtagcccac accagagtca gtttctgaat gtgacagcaa ctgaggatta
     1021 ttttgatcct aagacacagg agaataaaac aaatgtaaag gaattagagt actttgtcgg
     1081 acagagcttc gcgaatgcga tgtacaatgc ctgccgtgat gtggaggcgc cttccagtaa
     1141 cgagaaggcc ttaggactct tgtgtgggag ggatgcccgt gcctgcaatg ccaccaactg
     1201 gattgagtac atgttcaata aagacaacgg acaagcgcca tttaccatca ttcctgtgtt
     1261 ttcagatctt tcaatccttg ggatggagcc catgagaaat gccaccaaag gctgcaatga
     1321 gtctgtagat gaggtcacgg ggccatgtag ctgccaggac tgctccatcg tctgcggccc
     1381 caagccccag cccccacccc ctcctatgcc ctggaggatc tggggcttgg atgccatgta
     1441 tgtcatcatg tgggtcacct acgtggcatt tctgtttgtg ttttttggag cactgttggc
     1501 agtgtggtgc cacagaaggc ggtactttgt gtctgagtac actcccattg acagtaacat
     1561 cgccttttct gtgaatagca gtgacaaagg ggaagcctca tgctgtgacc cacttggtgc
     1621 agcatttgat gactgtctga ggcgcatgtt cacaaagtgg ggggctttct gtgtccgaaa
     1681 tcccacctgc atcattttct tctcattggc cttcatcact gtgtgctctt ctggcctggt
     1741 atttgtccag gtcaccacca atcctgtaga gctctggtca gcccctcaca gtcaggcccg
     1801 cttggaaaag gagtactttg acaagcactt tgggcctttc tttcgcacgg agcagcttat
     1861 catccaggcc cccaacacca gtgttcatat ctacgaaccg taccccgcag gagccgatgt
     1921 gcccttcggg cctccattga acaaagagat tctgcaccag gttctggact tacagatcgc
     1981 cattgaaagc atcaccgcat cttacaacaa tgaaaccgtg acactgcagg acatctgtgt
     2041 ggcccccctc tctccataca acaagaactg caccattatg agtgtgttaa attacttcca
     2101 gaacagccat gcggtgctgg acagccaagt aggcgacgac ttctatatct acgctgatta
     2161 ccacacacac tttctgtact gtgtacgggc tcccgcctcc ttgaatgata cgagtttgct
     2221 ccacggtcct tgcctgggta catttggagg accggtgttc ccgtggcttg tgttgggtgg
     2281 ctatgatgat cagaactaca acaatgccac cgcgcttgtg atcaccttcc ccgtgaataa
     2341 ttactacaat gacacagaga ggctccagag agcctgggcc tgggagaaag agtttattag
     2401 ttttgtgaaa aactacaaga atccaaatct gaccatttct ttcactgctg agcgaagcat
     2461 cgaagatgag ctcaatcggg aaagtaacag tgacgtgttc accgtcatca tcagctacgt
     2521 cgtgatgttt ctgtacattt ccctcgccct gggtcacatc cagagctgca gcaggctcct
     2581 ggtggattct aagatctcgc tgggcattgc ggggatcctg atcgtgctaa gctcggtggc
     2641 ctgctctctg ggcatcttca gctacatggg gatgccgctg accctcatcg tcattgaggt
     2701 catcccattc ctggtgctgg ctgtcggggt ggacaacatc ttcattctag tgcagaccta
     2761 ccagagagat gagcgtcttc aggaggaaac gctggatcag cagctgggca ggatccttgg
     2821 agaagtggcc ccgaccatgt tcctttcatc cttttctgag acctcagcat ttttctttgg
     2881 ggcgctgtcc tcgatgccag ctgtgcacac cttctctctg tttgcgggaa tggccgtcct
     2941 cattgacttc ctccttcaga ttacctgctt tgtgagcctg ttggggttag atattaagag
     3001 gcaagagaaa aaccatctgg acatcctgtg ctgtgtcaga ggcgctgacg acggacaagg
     3061 tagccacgcc tccgaaagct acctgtttcg cttcttcaaa aactactttg ccccccttct
     3121 gctgaaggac tggctgaggc caattgtggt agcggtgttt gtgggcgttc tgtcattcag
     3181 tgttgcggtg gtgaacaaag tagacatcgg gttggatcag tctctctcaa tgccaaacga
     3241 ttcgtatgtg attgactatt tcaaatcact cgctcagtac ctgcactcgg gcccacccgt
     3301 gtactttgtc ctggaggaag gctataacta cagttcacgc aaagggcaga acatggtgtg
     3361 cggcggcatg ggctgtgaca atgactccct ggtgcagcag atatttaacg cagctgagct
     3421 ggacacctac acccgagtag gcttcgcccc ctcgtcctgg atcgatgact actttgactg
     3481 ggtctcgcca cagtcgtcct gctgcagact ctacaacgtc actcaccagt tctgcaatgc
     3541 ttctgtgatg gacccaacct gtgtccgctg cagacctctg actccagagg gtaaacagag
     3601 gcctcagggg aaagaattca tgaaattcct gcccatgttc ctttctgata accccaaccc
     3661 caagtgcggc aaagggggac atgctgctta cggttcagct gttaacattg tgggagatga
     3721 cacttacatt ggggccactt acttcatgac ctaccacacc atacttaaga cctccgctga
     3781 ctatactgat gccatgaaga aagctcggct aatagccagt aacatcacgg aaaccatgcg
     3841 ttctaagggg agtgactacc gcgtattccc ttacagtgtg ttctacgtct tctatgaaca
     3901 gtacctgacc attattgatg acaccatctt taacctcagt gtgtctctgg gctccatatt
     3961 tctggtgacc ttggtggttc tgggctgtga gctgtggtct gcggtcatca tgtgtatcac
     4021 catagccatg atcctggtca acatgttcgg tgtcatgtgg ctgtggggca tcagtctgaa
     4081 tgcggtctcc ttggtcaact tggtgatgag ctgcggcatt tctgtggagt tctgcagcca
     4141 tataacgaga gcattcacca tgagtaccaa aggaagccga gtgagccggg cggaagaggc
     4201 actggcccac atgggtagtt ctgtattcag tggaatcaca cttacgaaat ttggagggat
     4261 cgtggtgtta gcctttgcca aatctcaaat ttttgagata ttttacttca ggatgtactt
     4321 agccatggtc ttactcggag ccactcatgg actaatattt cttcccgtct tactcagtta
     4381 catagggccg tcggtgaata aagctaaaag acacaccact tacgagcgct acagagggac
     4441 agagagagaa cggctcctca attttcacca ccaccaccac cacggtaccg cgggcccggg
     4501 atccatcgcc accatggtga gcaagggcga ggagctgttc accggggtgg tgcccatcct
     4561 ggtcgagctg gacggcgacg taaacggcca caagttcagc gtgtccggcg agggcgaggg
     4621 cgatgccacc tacggcaagc tgaccctgaa gttcatctgc accaccggca agctgcccgt
     4681 gccctggccc accctcgtga ccaccctgac ctacggcgtg cagtgcttca gccgctaccc
     4741 cgaccacatg aagcagcacg acttcttcaa gtccgccatg cccgaaggct acgtccagga
     4801 gcgcaccatc ttcttcaagg acgacggcaa ctacaagacc cgcgccgagg tgaagttcga
     4861 gggcgacacc ctggtgaacc gcatcgagct gaagggcatc gacttcaagg aggacggcaa
     4921 catcctgggg cacaagctgg agtacaacta caacagccac aacgtctata tcatggccga
     4981 caagcagaag aacggcatca aggtgaactt caagatccgc cacaacatcg aggacggcag
     5041 cgtgcagctc gccgaccact accagcagaa cacccccatc ggcgacggcc ccgtgctgct
     5101 gcccgacaac cactacctga gcacccagtc cgccctgagc aaagacccca acgagaagcg
     5161 cgatcacatg gtcctgctgg agttcgtgac cgccgccggg atcactctcg gcatggacga
     5221 gctgtacaag taaagcggcc gcgactctag atcataatca gccataccac atttgtagag
     5281 gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca taaaatgaat
     5341 gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata aagcaatagc
     5401 atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa
     5461 ctcatcaatg tatcttaagg cgtaaattgt aagcgttaat attttgttaa aattcgcgtt
     5521 aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca aaatccctta
     5581 taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga acaagagtcc
     5641 actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc agggcgatgg
     5701 cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc gtaaagcact
     5761 aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc cggcgaacgt
     5821 ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg caagtgtagc
     5881 ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac agggcgcgtc
     5941 aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt tctaaataca
     6001 ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat aatattgaaa
     6061 aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt agggtgtgga
     6121 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     6181 accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag catgcatctc
     6241 aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct aactccgccc
     6301 agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc agaggccgag
     6361 gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg aggcctaggc
     6421 ttttgcaaag atcgatcaag agacaggatg aggatcgttt cgcatgattg aacaagatgg
     6481 attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg actgggcaca
     6541 acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg ggcgcccggt
     6601 tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg aggcagcgcg
     6661 gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg ttgtcactga
     6721 agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc tgtcatctca
     6781 ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc tgcatacgct
     6841 tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc gagcacgtac
     6901 tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc aggggctcgc
     6961 gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg atctcgtcgt
     7021 gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct tttctggatt
     7081 catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt tggctacccg
     7141 tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc tttacggtat
     7201 cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt tcttctgagc
     7261 gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc acgagatttc
     7321 gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg ggacgccggc
     7381 tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc tagggggagg
     7441 ctaactgaaa cacggaagga gacaataccg gaaggaaccc gcgctatgac ggcaataaaa
     7501 agacagaata aaacgcacgg tgttgggtcg tttgttcata aacgcggggt tcggtcccag
     7561 ggctggcact ctgtcgatac cccaccgaga ccccattggg gccaatacgc ccgcgtttct
     7621 tccttttccc caccccaccc cccaagttcg ggtgaaggcc cagggctcgc agccaacgtc
     7681 ggggcggcag gccctgccat agcctcaggt tactcatata tactttagat tgatttaaaa
     7741 cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct catgaccaaa
     7801 atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa gatcaaagga
     7861 tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa aaaaccaccg
     7921 ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc gaaggtaact
     7981 ggcttcagca gagcgcagat accaaatact gttcttctag tgtagccgta gttaggccac
     8041 cacttcaaga actctgtagc accgcctaca tacctcgctc tgctaatcct gttaccagtg
     8101 gctgctgcca gtggcgataa gtcgtgtctt accgggttgg actcaagacg atagttaccg
     8161 gataaggcgc agcggtcggg ctgaacgggg ggttcgtgca cacagcccag cttggagcga
     8221 acgacctaca ccgaactgag atacctacag cgtgagctat gagaaagcgc cacgcttccc
     8281 gaagggagaa aggcggacag gtatccggta agcggcaggg tcggaacagg agagcgcacg
     8341 agggagcttc cagggggaaa cgcctggtat ctttatagtc ctgtcgggtt tcgccacctc
     8401 tgacttgagc gtcgattttt gtgatgctcg tcaggggggc ggagcctatg gaaaaacgcc
     8461 agcaacgcgg cctttttacg gttcctggcc ttttgctggc cttttgctca catgttcttt
     8521 cctgcgttat cccctgattc tgtggataac cgtattaccg ccatgcatta gttattaata
     8581 gtaatcaatt acggggtcat tagttcatag cccatatatg gagttccg
//
