LOCUS       sequence-378603-        6624 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses dominant-negative Vamp2 (mouse Vamp2 AA 1-94) plus 
            membrane-targeted EGFP in oligodendrocytes; AAV vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6624)
  AUTHORS   Lam M, Takeo K, Almeida RG, Cooper MH, Wu K, Iyer M, Kantarci H, 
            Zuchero JB
  TITLE     CNS myelination requires VAMP2/3-mediated membrane expansion in 
            oligodendrocytes.
  JOURNAL   Nat Commun. 2022 Sep 23;13(1):5583. doi: 10.1038/s41467-022-33200-4.
  PUBMED    36151203
REFERENCE   2  (bases 1 to 6624)
  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: "Nat 
            Commun."; date: "2022-09-23"; volume: "13(1):5583. doi"; pages: " 
            10.1038/s41467-022-33200-4"
COMMENT     Sequence Label: AAV_pMBP-dnVamp2-P2AT2A-EGFP-caax
FEATURES             Location/Qualifiers
     source          1..6624
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   11..136
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     CDS             2449..2505
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             2515..2568
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     CDS             join(2575..2577,2578..2580,2581..3291)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2575..2577,2578..2580,2581..3291)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(2620..2641)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2881..2900)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3228..3249
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    3381..3857
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3580..3599)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3897..4037
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3897..4026
                     /label=AAV2 ITR
     rep_origin      4112..4567
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4199..4218)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4409..4430
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4584..4603)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4703..4725
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4763..4781)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4849..4953
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4954..5022,5023..5814)
                     /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(5172..5191)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5985..6573
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6474..6493
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 gctcacatgt cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg
       61 ggcgaccttt ggtcgcccgg cctcagtgag cgagcgcgca gagagggagt ggccaactcc
      121 atcactaggg gttcctgcgg ccgcacgcgt gagctccttc ctgcttaggc cgtgtctgct
      181 ctgtcagtct ttcctctggc tctagtctgt ccatctgcag ggataacctc ccaagggccc
      241 tcttgttggc tgttcacctt ccgtctgtct tcatagcctc ctgatgagag agcgcgcaca
      301 cacaggctac ctgactgatg tgttgctgca ctgtggggtc cagtgacaca gctgtggagg
      361 tcaggaaccc agaattaagc tgtctgggag ccacaagttt caagagtcca tgctatggct
      421 cctacatgca ggtggtgcca gcagcattcc ttggtgtcct gtggctgtgg ttgcatctct
      481 gactctcctc cacctatcac attccttctc cagtaccagg cctctgtgtc tcagttttcc
      541 tctctcacaa gatccctagc ttcaggattg cacccacctc cactagccat gagcccgtcc
      601 tcactggatc acgtctgcaa agactctatt tccagttaaa gtcacagaca cgggcataga
      661 gagggggata ggatatgaaa atatcagttg ggggaggggc attcaattct tagctcacat
      721 tttcttcatc caatttcccc cactagaatg taagctttga gagaaaaggg accagatctt
      781 attcctcacc gtggctttaa cacttagaga aaatgcatcc cctctaatca ataagtcatc
      841 gacagtgggt agatggagga acggcagtgc gtagtaggat gcgtgcaagc atagtctcgt
      901 gcatgggtgc atagatcgct gggcaggtgg acaaggtggg ggtggataaa gaagtgggta
      961 gatgattgat gttaggtaaa tatcactggg tggacagatg ggtggtaggt ggatggatgg
     1021 ttagaatagt cagaagaggg atggattgat aaggtgaaca gatgataaat gggtgataga
     1081 ctggaagggt tgtcaaaaga ggataaggga agtgtgagct agccgtattt ctaaggtcag
     1141 taatagagtt gggagaagag gttaagttac atccatttaa acctcacacg aagctgagag
     1201 ggaatggact tgctgccgtt ggtgaggaaa gcgttgcatt tcccgtgtgc ttggttgtga
     1261 agtgctcagg tcccacatga agcagtcagg ttactgcggc ttacagagga gccagatcca
     1321 aatgccccga gtaagcacgt ccccgagcca gaggcctcca gcggaatccg ggagagggat
     1381 tgctcagtgc cctgcttccc tggactgtaa gctgcagaaa gatgtgggaa gtcctgttct
     1441 ccactgagaa cactaaaagc accttttgtc aaacgaccgc ttcacatctg gggcttgtgc
     1501 actggtggcc ttttaaacca gagacaaccc acaagatacc taacctgcgg ggctctctgg
     1561 tacagtgagc aactcaggaa atgctttggc ttgattgctg tgggctctca ggccatcgcc
     1621 ctctggagtg gttcttttaa tgagaacctg aagattggcc cctgagccat gtataccaag
     1681 caagctcaat ccaggttagc tccctctggt tggggcaagc taacgtgctc cttgggcccc
     1741 gcgcgtaact gtgcgtttta taggagacag ctagttcaag accccaggaa gaaagcggct
     1801 ttgtccccct ctaggcctcg tacaggccca cattcatatc tcattgttgt tgcaggggag
     1861 gcagatgcga tccagaacaa tgggacctcg gctgaggaca cggcggtgac agactccaag
     1921 cacacagcag acccaaagaa taactggcaa ggcgcccacc cagctgaccc agggaaccgc
     1981 ccccacttga tccgcctctt ttcccgagat gccccgggaa gggaggacaa caccttcaaa
     2041 gacaggccct cagagtccga cgagcttcag accatccaag aagaccccac agcagcttcc
     2101 ggaagctcga ataattctag gatccgaaca tcgattgaat tcgccaccat gtcggctacc
     2161 gctgccaccg tcccgcctgc cgccccggcc ggcgagggtg gcccccctgc acctcctcca
     2221 aaccttacta gtaacaggag actgcagcag acccaggccc aggtggatga ggtggtggac
     2281 atcatgaggg tgaatgtgga caaggtcctg gagcgggacc agaagttgtc ggagctggat
     2341 gaccgtgcag atgccctcca ggcaggggcc tcccagtttg aaacaagtgc agccaagctc
     2401 aagcgcaaat actggtggaa aaacctcaag accggtagcg gcggaagcgc tactaacttc
     2461 agcctgctga agcaggctgg agacgtggag gagaaccctg gacctggaag cggagagggc
     2521 agagggagtc tgctaacatg cggtgacgtc gaggagaatc ctggacctgc tagcatggtg
     2581 agcaagggcg aggagctgtt caccggggtg gtgcccatcc tggtcgagct ggacggcgac
     2641 gtaaacggcc acaagttcag cgtgtccggc gagggcgagg gcgatgccac ctacggcaag
     2701 ctgaccctga agttcatctg caccaccggc aagctgcccg tgccctggcc caccctcgtg
     2761 accaccctga cctacggcgt gcagtgcttc agccgctacc ccgaccacat gaagcagcac
     2821 gacttcttca agtccgccat gcccgaaggc tacgtccagg agcgcaccat cttcttcaag
     2881 gacgacggca actacaagac ccgcgccgag gtgaagttcg agggcgacac cctggtgaac
     2941 cgcatcgagc tgaagggcat cgacttcaag gaggacggca acatcctggg gcacaagctg
     3001 gagtacaact acaacagcca caacgtctat atcatggccg acaagcagaa gaacggcatc
     3061 aaggtgaact tcaagatccg ccacaacatc gaggacggca gcgtgcagct cgccgaccac
     3121 taccagcaga acacccccat cggcgacggc cccgtgctgc tgcccgacaa ccactacctg
     3181 agcacccagt ccgccctgag caaagacccc aacgagaagc gcgatcacat ggtcctgctg
     3241 gagttcgtga ccgccgccgg gatcactctc ggcatggacg agctgtacaa gtccggactc
     3301 agatctaagc tgaaccctcc tgatgagagt ggccccggct gcatgagctg caagtgtgtg
     3361 ctctcctgat aaagatctac gggtggcatc cctgtgaccc ctccccagtg cctctcctgg
     3421 ccctggaagt tgccactcca gtgcccacca gccttgtcct aataaaatta agttgcatca
     3481 ttttgtctga ctaggtgtcc ttctataata ttatggggtg gaggggggtg gtatggagca
     3541 aggggcaagt tgggaagaca acctgtaggg cctgcggggt ctattgggaa ccaagctgga
     3601 gtgcagtggc acaatcttgg ctcactgcaa tctccgcctc ctgggttcaa gcgattctcc
     3661 tgcctcagcc tcccgagttg ttgggattcc aggcatgcat gaccaggctc agctaatttt
     3721 tgtttttttg gtagagacgg ggtttcacca tattggccag gctggtctcc aactcctaat
     3781 ctcaggtgat ctacccacct tggcctccca aattgctggg attacaggcg tgaaccactg
     3841 ctcccttccc tgtccttctg attttgtagg taaccacgtg cggaccgagc ggccgcagga
     3901 acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg
     3961 gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc
     4021 gcgcagctgc ctgcaggggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     4081 ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct gtagcggcgc attaagcgcg
     4141 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgcctt agcgcccgct
     4201 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     4261 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     4321 cttgatttgg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     4381 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     4441 aactctatct cgggctattc ttttgattta taagggattt tgccgatttc ggtctattgg
     4501 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgttt
     4561 acaattttat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc
     4621 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
     4681 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca
     4741 ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg
     4801 ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
     4861 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     4921 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
     4981 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     5041 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     5101 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     5161 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
     5221 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     5281 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     5341 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     5401 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     5461 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
     5521 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     5581 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     5641 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     5701 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     5761 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     5821 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     5881 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     5941 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     6001 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     6061 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     6121 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     6181 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     6241 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     6301 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     6361 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     6421 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     6481 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     6541 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     6601 ttcctggcct tttgctggcc tttt
//
