LOCUS       sequence-220228-        5829 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV expression of nuclear localized GFP with SV40 NLS.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5829)
  AUTHORS   Challis RC, Ravindra Kumar S, Chan KY, Challis C, Beadle K, Jang MJ,
            Kim HM, Rajendran PS, Tompkins JD, Shivkumar K, Deverman BE, 
            Gradinaru V
  TITLE     Systemic AAV vectors for widespread and targeted gene delivery in 
            rodents.
  JOURNAL   Nat Protoc. 2019 Feb;14(2):379-414. doi: 10.1038/s41596-018-0097-3.
  PUBMED    30626963
REFERENCE   2  (bases 1 to 5829)
  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"; journalName: 
            "Nat Protoc"; date: "2019-02"; volume: "14"; issue: "2"; pages: 
            "379-414"
COMMENT     Sequence Label: pAAV-CAG-NLS-GFP
FEATURES             Location/Qualifiers
     source          1..5829
                     /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        286..589
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     982..1004
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1066..1085
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     CDS             1182..1202
                     /codon_start=1
                     /product="nuclear localization signal of SV40 (simian virus
                     40) large T antigen"
                     /label=SV40 NLS
                     /translation="PKKKRKV"
     CDS             join(1212..1214,1215..1217,1218..1931)
                     /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(1212..1214,1215..1217,1218..1928)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(1257..1278)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1518..1537)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1865..1886
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    1956..2544
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2009..2029)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2427..2438)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2576..3052
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2775..2794)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3092..3232
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3092..3221
                     /label=AAV2 ITR
     rep_origin      3307..3762
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3394..3413)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3604..3625
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3779..3798)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3898..3920
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3958..3976)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4044..4148
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4149..4217,4218..5009)
                     /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(4367..4386)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5180..5768
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5669..5688
                     /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 gcgtaacact agttattaat agtaatcaat tagagggagt
      181 ggccaactcc atcactaggg gttcctgcgg ccgcacgcgt aacactagtt attaatagta
      241 atcaattacg gggtcattag ttcatagccc atatatggag ttccgcgtta cataacttac
      301 ggtaaatggc ccgcctggct gaccgcccaa cgacccccgc ccattgacgt caataatgac
      361 gtatgttccc atagtaacgc caatagggac tttccattga cgtcaatggg tggagtattt
      421 acggtaaact gcccacttgg cagtacatca agtgtatcat atgccaagta cgccccctat
      481 tgacgtcaat gacggtaaat ggcccgcctg gcattatgcc cagtacatga ccttatggga
      541 ctttcctact tggcagtaca tctacgtatt agtcatcgct attaccatgg tcgaggtgag
      601 ccccacgttc tgcttcactc tccccatctc ccccccctcc ccacccccaa ttttgtattt
      661 atttattttt taattatttt gtgcagcgat gggggcgggg gggggggggg ggcgcgcgcc
      721 gggggggggg gggggggggg gggggggggg gggggcgggg cggcgcggtg cggcgggctg
      781 taattagcgc ttggtttaat gacggcttgt ttcttttctg tggctgcgtg aaagccttga
      841 ggggctccgg gagcgccggc aggaaggaaa tgggcgggga gggccttcgt gcgtcgccgc
      901 gccgccgtcc ccttctccct ctccagcctc ggggctgtcc gcggggggac ggctgccttc
      961 gggggggacg gggcagggcg gggttcggct tctggcgtgt gaccggcggc tctagagcct
     1021 ctgctaacca tgttcatgcc ttcttctttt tcctacagct cctgggcaac gtgctggtta
     1081 ttgtgctgtc tcatcatttt ggcaaagaat tgcggcccaa cggtaccgcc accatgtcta
     1141 gtgatgatga ggctactgct gactctcaac attctactcc tccaaaaaag aagagaaagg
     1201 tagaagaccc catggtgagc aagggcgagg agctgttcac cggggtggtg cccatcctgg
     1261 tcgagctgga cggcgacgta aacggccaca agttcagcgt gtccggcgag ggcgagggcg
     1321 atgccaccta cggcaagctg accctgaagt tcatctgcac caccggcaag ctgcccgtgc
     1381 cctggcccac cctcgtgacc accctgacct acggcgtgca gtgcttcagc cgctaccccg
     1441 accacatgaa gcagcacgac ttcttcaagt ccgccatgcc cgaaggctac gtccaggagc
     1501 gcaccatctt cttcaaggac gacggcaact acaagacccg cgccgaggtg aagttcgagg
     1561 gcgacaccct ggtgaaccgc atcgagctga agggcatcga cttcaaggag gacggcaaca
     1621 tcctggggca caagctggag tacaactaca acagccacaa cgtctatatc atggccgaca
     1681 agcagaagaa cggcatcaag gtgaacttca agatccgcca caacatcgag gacggcagcg
     1741 tgcagctcgc cgaccactac cagcagaaca cccccatcgg cgacggcccc gtgctgctgc
     1801 ccgacaacca ctacctgagc acccagtccg ccctgagcaa agaccccaac gagaagcgcg
     1861 atcacatggt cctgctggag ttcgtgaccg ccgccgggat cactctcggc atggacgagc
     1921 tgtacaagta agaattcgat atcaagctta tcgataatca acctctggat tacaaaattt
     1981 gtgaaagatt gactggtatt cttaactatg ttgctccttt tacgctatgt ggatacgctg
     2041 ctttaatgcc tttgtatcat gctattgctt cccgtatggc tttcattttc tcctccttgt
     2101 ataaatcctg gttgctgtct ctttatgagg agttgtggcc cgttgtcagg caacgtggcg
     2161 tggtgtgcac tgtgtttgct gacgcaaccc ccactggttg gggcattgcc accacctgtc
     2221 agctcctttc cgggactttc gctttccccc tccctattgc cacggcggaa ctcatcgccg
     2281 cctgccttgc ccgctgctgg acaggggctc ggctgttggg cactgacaat tccgtggtgt
     2341 tgtcggggaa atcatcgtcc tttccttggc tgctcgccta tgttgccacc tggattctgc
     2401 gcgggacgtc cttctgctac gtcccttcgg ccctcaatcc agcggacctt ccttcccgcg
     2461 gcctgctgcc ggctctgcgg cctcttccgc gtcttcgcct tcgccctcag acgagtcgga
     2521 tctccctttg ggccgcctcc ccgcatcgat accgagcgct gctcgagaga tctacgggtg
     2581 gcatccctgt gacccctccc cagtgcctct cctggccctg gaagttgcca ctccagtgcc
     2641 caccagcctt gtcctaataa aattaagttg catcattttg tctgactagg tgtccttcta
     2701 taatattatg gggtggaggg gggtggtatg gagcaagggg caagttggga agacaacctg
     2761 tagggcctgc ggggtctatt gggaaccaag ctggagtgca gtggcacaat cttggctcac
     2821 tgcaatctcc gcctcctggg ttcaagcgat tctcctgcct cagcctcccg agttgttggg
     2881 attccaggca tgcatgacca ggctcagcta atttttgttt ttttggtaga gacggggttt
     2941 caccatattg gccaggctgg tctccaactc ctaatctcag gtgatctacc caccttggcc
     3001 tcccaaattg ctgggattac aggcgtgaac cactgctccc ttccctgtcc ttctgatttt
     3061 gtaggtaacc acgtgcggac cgagcggccg caggaacccc tagtgatgga gttggccact
     3121 ccctctctgc gcgctcgctc gctcactgag gccgggcgac caaaggtcgc ccgacgcccg
     3181 ggctttgccc gggcggcctc agtgagcgag cgagcgcgca gctgcctgca ggggcgcctg
     3241 atgcggtatt ttctccttac gcatctgtgc ggtatttcac accgcatacg tcaaagcaac
     3301 catagtacgc gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg
     3361 tgaccgctac acttgccagc gccttagcgc ccgctccttt cgctttcttc ccttcctttc
     3421 tcgccacgtt cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc
     3481 gatttagtgc tttacggcac ctcgacccca aaaaacttga tttgggtgat ggttcacgta
     3541 gtgggccatc gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta
     3601 atagtggact cttgttccaa actggaacaa cactcaactc tatctcgggc tattcttttg
     3661 atttataagg gattttgccg atttcggtct attggttaaa aaatgagctg atttaacaaa
     3721 aatttaacgc gaattttaac aaaatattaa cgtttacaat tttatggtgc actctcagta
     3781 caatctgctc tgatgccgca tagttaagcc agccccgaca cccgccaaca cccgctgacg
     3841 cgccctgacg ggcttgtctg ctcccggcat ccgcttacag acaagctgtg accgtctccg
     3901 ggagctgcat gtgtcagagg ttttcaccgt catcaccgaa acgcgcgaga cgaaagggcc
     3961 tcgtgatacg cctattttta taggttaatg tcatgataat aatggtttct tagacgtcag
     4021 gtggcacttt tcggggaaat gtgcgcggaa cccctatttg tttatttttc taaatacatt
     4081 caaatatgta tccgctcatg agacaataac cctgataaat gcttcaataa tattgaaaaa
     4141 ggaagagtat gagtattcaa catttccgtg tcgcccttat tccctttttt gcggcatttt
     4201 gccttcctgt ttttgctcac ccagaaacgc tggtgaaagt aaaagatgct gaagatcagt
     4261 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
     4321 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
     4381 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
     4441 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
     4501 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
     4561 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
     4621 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
     4681 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
     4741 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
     4801 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
     4861 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
     4921 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
     4981 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
     5041 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
     5101 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
     5161 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
     5221 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     5281 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     5341 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     5401 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     5461 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     5521 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     5581 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     5641 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     5701 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     5761 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     5821 ctcacatgt
//
