LOCUS       sequence-454032-        6994 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  An adeno-associated viral vector to express GFP-iCre fusion protein 
            under the human Nestin 2nd intron enhancer..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6994)
  TITLE     Ribeiro Gomes plasmids 2024
REFERENCE   2  (bases 1 to 6994)
  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"
COMMENT     Sequence Label: pOTTC2234 - pAAV Nestin GFP-iCre
FEATURES             Location/Qualifiers
     source          1..6994
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        2093..2155
                     /label=Mini-TK promoter
                     /note="minimal herpes simplex virus (HSV) thymidine kinase 
                     promoter"
     intron          2212..2308
                     /label=SV40 intron
                     /note="modified SV40 intron with splice donor and acceptor 
                     sites"
     regulatory      2378..2387
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(2384..2386,2387..2389,2390..3100)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(2384..2386,2387..2389,2390..3100)
                     /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(2429..2450)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2690..2709)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3037..3058
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             3107..3127
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     repeat_region   4257..4397
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4257..4386
                     /label=AAV2 ITR
     rep_origin      4472..4927
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4559..4578)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4769..4790
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4944..4963)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5063..5085
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5123..5141)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5209..5313
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5314..5382,5383..6174)
                     /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(5532..5551)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6345..6933
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6834..6853
                     /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 gcgtggagca ggagaaacag ggcctacaga gccagatcgc
      181 tcaggtcctg gaaggtcggc agcagctggc gcacctcaag atgtccctca gcctggaggt
      241 ggccacgtac aggtacgaga cgagccctgt ccacccaggc acacagaggc acacacacat
      301 gctggggcag actacgtggc tgaaggggac cagaggtgta ggtgatggca actctgctgc
      361 cactgggttc tgtgactcta ggctggtcac ttccacaccg gaacatgtcc tgcctgaaac
      421 cgtctgctcc cccataggct ggagacctga gcaatgtcct gtgcccagga aatcccccca
      481 ggagagatga tattgccgag cgggacagcc tctggagagg ggctgggccc gtgtggggca
      541 actgcatcca gacccttccc ccaaggggca aagctccaac tctatgtgtg cccgcaccca
      601 gggggtcatg tcctttggca gccacggaaa ggaggcattg caggcatggg gtggctgcac
      661 tgacttgcca cctgacctgg gcctcagcct cgtctagggg ctgagggctg ggctggggga
      721 cttccaaggc ctcttctagc ctggatgtca tggggcgaga ctcctctgat gcccctagtg
      781 cccttggtcc tgcccagcct ttcattgtcc acattttctg ttatcctaga aatggctgag
      841 accaaggagg ggtggcttct cacttgctat gtggccttgg gcaaagcccg cagtctcttc
      901 tgggaaactg gtcctgttgg ctgaggggcc attgatgggg cctgaggaag gggctctgtg
      961 gctggtgagg ggccacatga gatccaggaa aggaggagag gcaaggccca gagatggaag
     1021 gtcagtgagg ctctggggag gacgtggagg agaggggctc caccgccagg gaagagaagc
     1081 ccagaaggaa ggccgcagca ctggcaagga cagaggtgag gctacagcga gtgctaacca
     1141 gactcagaga tcatggagca aagcagagag gagcagctgc tcatcctaag ttattgaatt
     1201 gattggtccc tgctcccctc tttggttcaa ggggcgctga ggaaggggcc tggacacaaa
     1261 gacctctgtg ccccgaagag cttgggatcc agagtgctta gagatcctga gatgagggcc
     1321 ctgaccccca gtgaagccac cgaggggaag ggtctggtgg gccccgagga atttaactgc
     1381 tattttttga ggcatgaaaa ggatttggag agggggagct gaattcattt gcttttgtct
     1441 gtcgctagct ctgggggcca ccggggaagg aggagggagc ccatcccatg ggaacggcct
     1501 gagaattccc acttccccag aatcctctcc ttctcagacc ctccagatgg tggcttggac
     1561 aatggcagca attagcatga gaatcggtcc cccacacaga ggatgaggtc attggcctgg
     1621 ggtgggagga ggaggctagc ctaagccttg tgtgggtgtg tgtagggcaa tgtccgactg
     1681 tgtgggaatc tgcttctatg tctgagctct gctgtgtatc ccaaatacag tgtgactttg
     1741 tgtgaccaag tgtgtgatgt ctgtgggtat gtgactgtgt gatttcaccc tgagtgtggc
     1801 ttttgtgtga cagtgtcaaa tgaagaggaa gcctgcaggg cctggatggt gcaagctcag
     1861 cctgagttgt ccccagctgg cttgctgcct gcaggcctgg gggcagaggg ctgaccccac
     1921 tttccctctg tcctctccag gaccctcctg gaggctgaga actcccggct gcaaacacct
     1981 ggcggtggct ccaagacttg tcgactctag aggatccggc cccgcccagc gtcttgtcat
     2041 tggcgaattc gaacacgcag atgcagtcgg ggcggcgcgg tccgaggtcc acttcgcata
     2101 ttaaggtgac gcgtgtggcc tcgaacaccg agcgaccctg cagcgacccg cttaacagcg
     2161 tcaacagcgt gccgcagatc tcgaggaact gaaaaaccag aaagttaact ggtaagttta
     2221 gtctttttgt cttttatttc aggtcccgga tccggtggtg gtgcaaatca aagaactgct
     2281 cctcagtgga tgttgccttt acttctaggc ctgtacggaa gtgttacttc tgctctaaaa
     2341 gctgcggaat tgtacccgcg gccgggatcc accggtcgcc accatggtga gcaagggcga
     2401 ggagctgttc accggggtgg tgcccatcct ggtcgagctg gacggcgacg taaacggcca
     2461 caagttcagc gtgtccggcg agggcgaggg cgatgccacc tacggcaagc tgaccctgaa
     2521 gttcatctgc accaccggca agctgcccgt gccctggccc accctcgtga ccaccctgac
     2581 ctacggcgtg cagtgcttca gccgctaccc cgaccacatg aagcagcacg acttcttcaa
     2641 gtccgccatg cccgaaggct acgtccagga gcgcaccatc ttcttcaagg acgacggcaa
     2701 ctacaagacc cgcgccgagg tgaagttcga gggcgacacc ctggtgaacc gcatcgagct
     2761 gaagggcatc gacttcaagg aggacggcaa catcctgggg cacaagctgg agtacaacta
     2821 caacagccac aacgtctata tcatggccga caagcagaag aacggcatca aggtgaactt
     2881 caagatccgc cacaacatcg aggacggcag cgtgcagctc gccgaccact accagcagaa
     2941 cacccccatc ggcgacggcc ccgtgctgct gcccgacaac cactacctga gcacccagtc
     3001 cgccctgagc aaagacccca acgagaagcg cgatcacatg gtcctgctgg agttcgtgac
     3061 cgccgccggg atcactctcg gcatggacga gctgtacaag ggaggaccca agaagaagag
     3121 gaaagtctcc aacctgctga ctgtgcacca aaacctgcct gccctccctg tggatgccac
     3181 ctctgatgaa gtcaggaaga acctgatgga catgttcagg gacaggcagg ccttctctga
     3241 acacacctgg aagatgctcc tgtctgtgtg cagatcctgg gctgcctggt gcaagctgaa
     3301 caacaggaaa tggttccctg ctgaacctga ggatgtgagg gactacctcc tgtacctgca
     3361 agccagaggc ctggctgtga agaccatcca acagcacctg ggccagctca acatgctgca
     3421 caggagatct ggcctgcctc gcccttctga ctccaatgct gtgtccctgg tgatgaggag
     3481 aatcagaaag gagaatgtgg atgctgggga gagagccaag caggccctgg cctttgaacg
     3541 cactgacttt gaccaagtca gatccctgat ggagaactct gacagatgcc aggacatcag
     3601 gaacctggcc ttcctgggca ttgcctacaa caccctgctg cgcattgccg aaattgccag
     3661 aatcagagtg aaggacatct cccgcaccga tggtgggaga atgctgatcc acattggcag
     3721 gaccaagacc ctggtgtcca cagctggtgt ggagaaggcc ctgtccctgg gggttaccaa
     3781 gctggtggag agatggatct ctgtgtctgg tgtggctgat gaccccaaca actacctgtt
     3841 ctgccgggtc agaaagaatg gtgtggctgc cccttctgcc acctcccaac tgtccacccg
     3901 ggccctggaa gggatctttg aggccaccca ccgcctgatc tatggtgcca aggatgactc
     3961 tgggcagaga tacctggcct ggtctggcca ctctgccaga gtgggtgctg ccagggacat
     4021 ggccagggct ggtgtgtcca tccctgaaat catgcaggct ggtggctgga ccaatgtgaa
     4081 cattgtgatg aactacatca gaaacctgga ctctgagact ggggccatgg tgaggctgct
     4141 cgaggatggg gactgactcg agggcaataa aaagacagaa taaaacgcac gggtgttggg
     4201 tcgtttgttc ataaacgcgg ggttcggtgg taaccacgtg cggaccgagc ggccgcagga
     4261 acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg
     4321 gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc
     4381 gcgcagctgc ctgcaggggc gcctgatgcg gtattttctc cttacgcatc tgtgcggtat
     4441 ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct gtagcggcgc attaagcgcg
     4501 gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg ccagcgcctt agcgcccgct
     4561 cctttcgctt tcttcccttc ctttctcgcc acgttcgccg gctttccccg tcaagctcta
     4621 aatcgggggc tccctttagg gttccgattt agtgctttac ggcacctcga ccccaaaaaa
     4681 cttgatttgg gtgatggttc acgtagtggg ccatcgccct gatagacggt ttttcgccct
     4741 ttgacgttgg agtccacgtt ctttaatagt ggactcttgt tccaaactgg aacaacactc
     4801 aactctatct cgggctattc ttttgattta taagggattt tgccgatttc ggtctattgg
     4861 ttaaaaaatg agctgattta acaaaaattt aacgcgaatt ttaacaaaat attaacgttt
     4921 acaattttat ggtgcactct cagtacaatc tgctctgatg ccgcatagtt aagccagccc
     4981 cgacacccgc caacacccgc tgacgcgccc tgacgggctt gtctgctccc ggcatccgct
     5041 tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc agaggttttc accgtcatca
     5101 ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat ttttataggt taatgtcatg
     5161 ataataatgg tttcttagac gtcaggtggc acttttcggg gaaatgtgcg cggaacccct
     5221 atttgtttat ttttctaaat acattcaaat atgtatccgc tcatgagaca ataaccctga
     5281 taaatgcttc aataatattg aaaaaggaag agtatgagta ttcaacattt ccgtgtcgcc
     5341 cttattccct tttttgcggc attttgcctt cctgtttttg ctcacccaga aacgctggtg
     5401 aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg gttacatcga actggatctc
     5461 aacagcggta agatccttga gagttttcgc cccgaagaac gttttccaat gatgagcact
     5521 tttaaagttc tgctatgtgg cgcggtatta tcccgtattg acgccgggca agagcaactc
     5581 ggtcgccgca tacactattc tcagaatgac ttggttgagt actcaccagt cacagaaaag
     5641 catcttacgg atggcatgac agtaagagaa ttatgcagtg ctgccataac catgagtgat
     5701 aacactgcgg ccaacttact tctgacaacg atcggaggac cgaaggagct aaccgctttt
     5761 ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt gggaaccgga gctgaatgaa
     5821 gccataccaa acgacgagcg tgacaccacg atgcctgtag caatggcaac aacgttgcgc
     5881 aaactattaa ctggcgaact acttactcta gcttcccggc aacaattaat agactggatg
     5941 gaggcggata aagttgcagg accacttctg cgctcggccc ttccggctgg ctggtttatt
     6001 gctgataaat ctggagccgg tgagcgtggg tctcgcggta tcattgcagc actggggcca
     6061 gatggtaagc cctcccgtat cgtagttatc tacacgacgg ggagtcaggc aactatggat
     6121 gaacgaaata gacagatcgc tgagataggt gcctcactga ttaagcattg gtaactgtca
     6181 gaccaagttt actcatatat actttagatt gatttaaaac ttcattttta atttaaaagg
     6241 atctaggtga agatcctttt tgataatctc atgaccaaaa tcccttaacg tgagttttcg
     6301 ttccactgag cgtcagaccc cgtagaaaag atcaaaggat cttcttgaga tccttttttt
     6361 ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc taccagcggt ggtttgtttg
     6421 ccggatcaag agctaccaac tctttttccg aaggtaactg gcttcagcag agcgcagata
     6481 ccaaatactg ttcttctagt gtagccgtag ttaggccacc acttcaagaa ctctgtagca
     6541 ccgcctacat acctcgctct gctaatcctg ttaccagtgg ctgctgccag tggcgataag
     6601 tcgtgtctta ccgggttgga ctcaagacga tagttaccgg ataaggcgca gcggtcgggc
     6661 tgaacggggg gttcgtgcac acagcccagc ttggagcgaa cgacctacac cgaactgaga
     6721 tacctacagc gtgagctatg agaaagcgcc acgcttcccg aagggagaaa ggcggacagg
     6781 tatccggtaa gcggcagggt cggaacagga gagcgcacga gggagcttcc agggggaaac
     6841 gcctggtatc tttatagtcc tgtcgggttt cgccacctct gacttgagcg tcgatttttg
     6901 tgatgctcgt caggggggcg gagcctatgg aaaaacgcca gcaacgcggc ctttttacgg
     6961 ttcctggcct tttgctggcc ttttgctcac atgt
//
