LOCUS       Exported                5859 bp ds-DNA     circular SYN 07-AUG-2025
DEFINITION  AAV vector expressing Myc-tagged SpCas9 under the EFS promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5859)
  AUTHORS   Nishiyama J, Mikuni T, Yasuda R
  TITLE     Virus-Mediated Genome Editing via Homology-Directed Repair in 
            Mitotic and Postmitotic Cells in Mammalian Brain.
  JOURNAL   Neuron. 2017 Oct 18. pii: S0896-6273(17)30933-9. doi: 
            10.1016/j.neuron.2017.10.004.
  PUBMED    29056297
REFERENCE   2  (bases 1 to 5859)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Jan 10, 2018 from SnapGene Server 1.1.58
            http://www.snapgene.com
REFERENCE   3  (bases 1 to 5859)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Thursday, Aug 7, 2025 from SnapGene 5.0.8
            https://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..5859
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     source          join(3711..5859,1..787)
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     rep_origin      1..589
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of  
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   651..780
                     /label=ITR
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     misc_feature    join(788..815,2882..2899,2904..2931)
                     /label=phagemid pBluescriptII SK(+) [Split]
                     /note="phagemid pBluescriptII SK(+) [Split]"
     N_region        complement(join(827..843,1881..2013))
                     /label=attL1 [Split]
                     /note="attL1 [Split]"
     promoter        join(907..1449,1881..1984)
                     /label=GFA-ABC1D
                     /note="GFAP promoter of 681 bp from Dr M. Brenner 
                     corresponding to GFAP promoter sequences -1757 to -1256 / 
                     -132 to +47 [Split]"
     misc_feature    1485..1877
                     /label=(B)3
                     /note="/(B)3="
     gene            2052..2801
                     /label=GFP
                     /note="/GFP="
     N_region        2872..2881
                     /label=attL2 [Split]
                     /note="attL2 [Split]"
     repeat_region   2882..3710
     repeat_region   2882..2931
     repeat_region   2882..2931
     repeat_region   2882..2931
                     /label=RFA
     gene            2932
                     /label=EGFP=
                     /note="EGFP="
     misc_feature    3564..3668
                     /label=miR124T(N)
                     /note="miR124T Naldini"
     polyA_signal    3716..3764
                     /label=synthetic polyadenylation signal
                     /note="synthetic polyadenylation signal"
     repeat_region   3772..3912
                     /label=ITR
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus  
                     serotype 2"
     repeat_region   3772..3901
                     /label=AAV2 ITR
     rep_origin      3987..4442
                     /label=f1 ori
                     /note="""f1 bacteriophage origin of replication"
                     /note="arrow  indicates direction of (+) strand 
                     synthesis"""
     primer_bind     complement(4074..4093)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4284..4305
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4459..4478)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable  
                     marker"
     primer_bind     4578..4600
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4638..4656)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward  
                     primer"
     promoter        4724..4828
                     /gene="bla"
                     /label=AmpR promoter
     CDS             4829..5689
                     /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(5047..5066)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt cctgcaggca
      661 gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt ggtcgcccgg
      721 cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact aggggttcct
      781 gcggcctcta gtggatcccc cgggctgcag gaattatcac aagtttgtac aaaaaagcag
      841 gctccaccat gggaaccaat tcagtcgact ggatccggta ccgaattcga gctcggtacc
      901 agatctaaca tatcctggtg tggagtaggg gacgctgctc tgacagaggc tcgggggcct
      961 gagctggctc tgtgagctgg ggaggaggca gacagccagg ccttgtctgc aagcagacct
     1021 ggcagcattg ggctggccgc cccccagggc ctcctcttca tgcccagtga atgactcacc
     1081 ttggcacaga cacaatgttc ggggtgggca cagtgcctgc ttcccgccgc accccagccc
     1141 ccctcaaatg ccttccgaga agcccattga gcagggggct tgcattgcac cccagcctga
     1201 cagcctggca tcttgggata aaagcagcac agccccctag gggctgccct tgctgtgtgg
     1261 cgccaccggc ggtggagaac aaggctctat tcagcctgtg cccaggaaag gggatcaggg
     1321 gatgcccagg catggacagt gggtggcagg gggggagagg agggctgtct gcttcccaga
     1381 agtccaagga cacaaatggg tgaggggaga gctctcccca tagctgggct gcggcccaac
     1441 cccaccccct caggctatgc cagggggtgt tgccaggggc acccgatctt cctcttcatg
     1501 cccagtgaat gactcacctt ggcacagaca caatgttcgg ggtgggcaca gtgcctgctt
     1561 cccgccgcac cccagccccc ctcaaatgcc ttccgagaag cccattgagc aggcccgatc
     1621 ttcctcttca tgcccagtga atgactcacc ttggcacaga cacaatgttc ggggtgggca
     1681 cagtgcctgc ttcccgccgc accccagccc ccctcaaatg ccttccgaga agcccattga
     1741 gcaggcccga tcttcctctt catgcccagt gaatgactca ccttggcaca gacacaatgt
     1801 tcggggtggg cacagtgcct gcttcccgcc gcaccccagc ccccctcaaa tgccttccga
     1861 gaagcccatt gagcaggccc gggcatcgcc agtctagccc actccttcat aaagccctcg
     1921 catcccagga gcgagcagag ccagagcagg ttggagagga gacgcatcac ctccgctgct
     1981 cgcggggatc ctctagagtc gacggatccg gggaattcgc ggcccgcgtc ctagcgctac
     2041 cggtcgccac catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg
     2101 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg
     2161 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
     2221 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg
     2281 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
     2341 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg
     2401 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     2461 tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca
     2521 agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg
     2581 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     2641 ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg
     2701 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc
     2761 tgtacaagtc cggactcaga tctcgactag ctagtagcta gctagctagt cgagctcaag
     2821 cttcgaattc gatatcaagc ttatcgatac cgtcgacctc gagatatcta gacccagctt
     2881 tcttgtacaa agtggtgata attcgatatc aagcttatcg ataccgtcga cctcgaggga
     2941 attccgataa tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact
     3001 atgttgctcc ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg
     3061 cttcccgtat ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg
     3121 aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa
     3181 cccccactgg ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc
     3241 ccctccctat tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg
     3301 ctcggctgtt gggcactgac aattccgtgg tgttgtcggg gaagctgacg tcctttccat
     3361 ggctgctcgc ctgtgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt
     3421 cggccctcaa tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc
     3481 cgcgtcttcg ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc
     3541 gggaattcga gctcggtacc taatggcatt caccgcgtgc cttaattcga atggcattca
     3601 ccgcgtgcct taaacgcgtt ggcattcacc gcgtgcctta aatgcattgg cattcaccgc
     3661 gtgccttaag gtacctttaa gaccaatgac ttacaaggca gctgtagatc aattcaataa
     3721 aagatcttta ttttcattag atctgtgtgt tggttttttg tgtgcggccg caggaacccc
     3781 tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag gccgggcgac
     3841 caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag cgagcgcgca
     3901 gctgcctgca ggggcgcctg atgcggtatt ttctccttac gcatctgtgc ggtatttcac
     3961 accgcatacg tcaaagcaac catagtacgc gccctgtagc ggcgcattaa gcgcggcggg
     4021 tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccttagcgc ccgctccttt
     4081 cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg
     4141 ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca aaaaacttga
     4201 tttgggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc gccctttgac
     4261 gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa cactcaactc
     4321 tatctcgggc tattcttttg atttataagg gattttgccg atttcggtct attggttaaa
     4381 aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa cgtttacaat
     4441 tttatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc agccccgaca
     4501 cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat ccgcttacag
     4561 acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa
     4621 acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg tcatgataat
     4681 aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa cccctatttg
     4741 tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac cctgataaat
     4801 gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg tcgcccttat
     4861 tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt
     4921 aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg atctcaacag
     4981 cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga gcacttttaa
     5041 agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc aactcggtcg
     5101 ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag aaaagcatct
     5161 tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga gtgataacac
     5221 tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg cttttttgca
     5281 caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga atgaagccat
     5341 accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact
     5401 attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact ggatggaggc
     5461 ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt ttattgctga
     5521 taaatctgga gccggtgagc gtggaagccg cggtatcatt gcagcactgg ggccagatgg
     5581 taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta tggatgaacg
     5641 aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac tgtcagacca
     5701 agtttactca tatatacttt agattgattt aaaacttcat ttttaattta aaaggatcta
     5761 ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt tttcgttcca
     5821 ctgagcgtca gaccccgtag aaaagatcaa aggatcttc
//
