LOCUS       sequence-330043-        4907 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses nuclear EGFP from CMV promoter.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4907)
  AUTHORS   Maturana CJ, Verpeut JL, Kooshkbaghi M, Engel EA
  TITLE     Novel tool to quantify with single-cell resolution the number of 
            incoming AAV genomes co-expressed in the mouse nervous system.
  JOURNAL   Gene Ther. 2021 Jun 28. pii: 10.1038/s41434-021-00272-8. doi: 
            10.1038/s41434-021-00272-8.
  PUBMED    34176926
REFERENCE   2  (bases 1 to 4907)
  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: "Gene Ther. 
            2021 Jun 28. pii: 10.1038/s41434-021-00272-8. doi: 
            10.1038/s41434-021-00272-8."
COMMENT     Sequence Label: pAAV-CMV-EGFP-NLS
FEATURES             Location/Qualifiers
     source          1..4907
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR
     promoter        191..394
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     345..365
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    417..497
                     /label=MCS
                     /note="multiple cloning site"
     regulatory      499..508
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(505..507,508..510,511..1221)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             join(505..507,508..510,511..1221)
                     /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(550..571)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(811..830)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1158..1179
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             1237..1263
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             1264..1290
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     CDS             1291..1317
                     /codon_start=1
                     /product="nuclear localization signal of human c-Myc 
                     proto-oncogene (Dang and Lee, 1988)"
                     /label=c-myc NLS
                     /translation="PAAKRVKLD"
     misc_feature    1345..1933
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1398..1418)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1816..1827)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(1936..1952)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1937..1953)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    1982..2103
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1993..2012)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     2047..2066
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   2265..2394
                     /label=AAV2 ITR
     rep_origin      complement(2631..3219)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2711..2730)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(3390..4181,4182..4250))
                     /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     4013..4032
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4251..4355)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(4381..4836)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4518..4539)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     4730..4749
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 ctgcgcgctc gctcgctcac tgaggccgcc cgggcaaagc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct tgtagttaat gattaacccg ccatgctact tatctacgta gccatgctct
      181 aggaagatct gtgatgcggt tttggcagta catcaatggg cgtggatagc ggtttgactc
      241 acggggattt ccaagtctcc accccattga cgtcaatggg agtttgtttt ggcaccaaaa
      301 tcaacgggac tttccaaaat gtcgtaacaa ctccgcccca ttgacgcaaa tgggcggtag
      361 gcgtgtacgg tgggaggtct atataagcag agctggttta gtgaaccgtc agatccgcta
      421 gcgctaccgg actcagatct cgagctcaag cttcgaattc tgcagtcgac ggtaccgcgg
      481 gcccgggatc caccggtcgc caccatggtg agcaagggcg aggagctgtt caccggggtg
      541 gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc
      601 gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc
      661 aagctgcccg tgccctggcc caccctcgtg accaccctga cctacggcgt gcagtgcttc
      721 agccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc
      781 tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag
      841 gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag
      901 gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat
      961 atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc
     1021 gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc
     1081 cccgtgctgc tgcccgacaa ccactacctg agcacccagt ccgccctgag caaagacccc
     1141 aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc
     1201 ggcatggacg agctgtacaa gagatctcga gctgatcctg ctgccaaacg cgttaaacta
     1261 gaccctgctg ccaaacgcgt taaactagac cctgctgcca aacgcgttaa actagactaa
     1321 agcggccgcg acaagcttat cgataatcaa cctctggatt acaaaatttg tgaaagattg
     1381 actggtattc ttaactatgt tgctcctttt acgctatgtg gatacgctgc tttaatgcct
     1441 ttgtatcatg ctattgcttc ccgtatggct ttcattttct cctccttgta taaatcctgg
     1501 ttgctgtctc tttatgagga gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact
     1561 gtgtttgctg acgcaacccc cactggttgg ggcattgcca ccacctgtca gctcctttcc
     1621 gggactttcg ctttccccct ccctattgcc acggcggaac tcatcgccgc ctgccttgcc
     1681 cgctgctgga caggggctcg gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa
     1741 tcatcgtcct ttccttggct gctcgcctgt gttgccacct ggattctgcg cgggacgtcc
     1801 ttctgctacg tcccttcggc cctcaatcca gcggaccttc cttcccgcgg cctgctgccg
     1861 gctctgcggc ctcttccgcg tcttcgcctt cgccctcaga cgagtcggat ctccctttgg
     1921 gccgcctccc cgcatcgata ccgtcgacct cgacccgggc ggccgcttcg agcagacatg
     1981 ataagataca ttgatgagtt tggacaaacc acaactagaa tgcagtgaaa aaaatgcttt
     2041 atttgtgaaa tttgtgatgc tattgcttta tttgtaacca ttataagctg caataaacaa
     2101 gttaacaaca acaattgcat tcattttatg tttcaggttc agggggagat gtgggaggtt
     2161 ttttaaagca agtaaaacct ctacaaatgt ggtaaaatcg ataaggatct tcctagagca
     2221 tggctacgta gataagtagc atggcgggtt aatcattaac tacaaggaac ccctagtgat
     2281 ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt
     2341 cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagctgcat
     2401 taatgaatcg gccaacgcgc ggggagaggc ggtttgcgta ttgggcgctc ttccgcttcc
     2461 tcgctcactg actcgctgcg ctcggtcgtt cggctgcggc gagcggtatc agctcactca
     2521 aaggcggtaa tacggttatc cacagaatca ggggataacg caggaaagaa catgtgagca
     2581 aaaggccagc aaaaggccag gaaccgtaaa aaggccgcgt tgctggcgtt tttccatagg
     2641 ctccgccccc ctgacgagca tcacaaaaat cgacgctcaa gtcagaggtg gcgaaacccg
     2701 acaggactat aaagatacca ggcgtttccc cctggaagct ccctcgtgcg ctctcctgtt
     2761 ccgaccctgc cgcttaccgg atacctgtcc gcctttctcc cttcgggaag cgtggcgctt
     2821 tctcatagct cacgctgtag gtatctcagt tcggtgtagg tcgttcgctc caagctgggc
     2881 tgtgtgcacg aaccccccgt tcagcccgac cgctgcgcct tatccggtaa ctatcgtctt
     2941 gagtccaacc cggtaagaca cgacttatcg ccactggcag cagccactgg taacaggatt
     3001 agcagagcga ggtatgtagg cggtgctaca gagttcttga agtggtggcc taactacggc
     3061 tacactagaa gaacagtatt tggtatctgc gctctgctga agccagttac cttcggaaaa
     3121 agagttggta gctcttgatc cggcaaacaa accaccgctg gtagcggtgg tttttttgtt
     3181 tgcaagcagc agattacgcg cagaaaaaaa ggatctcaag aagatccttt gatcttttct
     3241 acggggtctg acgctcagtg gaacgaaaac tcacgttaag ggattttggt catgagatta
     3301 tcaaaaagga tcttcaccta gatcctttta aattaaaaat gaagttttaa atcaatctaa
     3361 agtatatatg agtaaacttg gtctgacagt taccaatgct taatcagtga ggcacctatc
     3421 tcagcgatct gtctatttcg ttcatccata gttgcctgac tccccgtcgt gtagataact
     3481 acgatacggg agggcttacc atctggcccc agtgctgcaa tgataccgcg agacccacgc
     3541 tcaccggctc cagatttatc agcaataaac cagccagccg gaagggccga gcgcagaagt
     3601 ggtcctgcaa ctttatccgc ctccatccag tctattaatt gttgccggga agctagagta
     3661 agtagttcgc cagttaatag tttgcgcaac gttgttgcca ttgctacagg catcgtggtg
     3721 tcacgctcgt cgtttggtat ggcttcattc agctccggtt cccaacgatc aaggcgagtt
     3781 acatgatccc ccatgttgtg caaaaaagcg gttagctcct tcggtcctcc gatcgttgtc
     3841 agaagtaagt tggccgcagt gttatcactc atggttatgg cagcactgca taattctctt
     3901 actgtcatgc catccgtaag atgcttttct gtgactggtg agtactcaac caagtcattc
     3961 tgagaatagt gtatgcggcg accgagttgc tcttgcccgg cgtcaatacg ggataatacc
     4021 gcgccacata gcagaacttt aaaagtgctc atcattggaa aacgttcttc ggggcgaaaa
     4081 ctctcaagga tcttaccgct gttgagatcc agttcgatgt aacccactcg tgcacccaac
     4141 tgatcttcag catcttttac tttcaccagc gtttctgggt gagcaaaaac aggaaggcaa
     4201 aatgccgcaa aaaagggaat aagggcgaca cggaaatgtt gaatactcat actcttcctt
     4261 tttcaatatt attgaagcat ttatcagggt tattgtctca tgagcggata catatttgaa
     4321 tgtatttaga aaaataaaca aataggggtt ccgcgcacat ttccccgaaa agtgccacct
     4381 aaattgtaag cgttaatatt ttgttaaaat tcgcgttaaa tttttgttaa atcagctcat
     4441 tttttaacca ataggccgaa atcggcaaaa tcccttataa atcaaaagaa tagaccgaga
     4501 tagggttgag tgttgttcca gtttggaaca agagtccact attaaagaac gtggactcca
     4561 acgtcaaagg gcgaaaaacc gtctatcagg gcgatggccc actacgtgaa ccatcaccct
     4621 aatcaagttt tttggggtcg aggtgccgta aagcactaaa tcggaaccct aaagggagcc
     4681 cccgatttag agcttgacgg ggaaagccgg cgaacgtggc gagaaaggaa gggaagaaag
     4741 cgaaaggagc gggcgctagg gcgctggcaa gtgtagcggt cacgctgcgc gtaaccacca
     4801 cacccgccgc gcttaatgcg ccgctacagg gcgcgtccca ttcgccattc aggctgcgca
     4861 actgttggga agggcgatcg gtgcgggcct cttcgctatt acgccag
//
