LOCUS       sequence-377185-        4574 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses EGFP in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 4574)
  AUTHORS   Sohn J, Takahashi M, Okamoto S, Ishida Y, Furuta T, Hioki H
  TITLE     A Single Vector Platform for High-Level Gene Transduction of Central
            Neurons: Adeno-Associated Virus Vector Equipped with the Tet-Off 
            System.
  JOURNAL   PLoS One. 2017 Jan 6;12(1):e0169611. doi: 
            10.1371/journal.pone.0169611. eCollection 2017.
  PUBMED    28060929
REFERENCE   2  (bases 1 to 4574)
  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: "PLoS One.";
            date: "2017-01-6"; volume: "12(1):e0169611. doi"; pages: " 
            10.1371/journal.pone.0169611. eCollection 2017"
COMMENT     Sequence Label: pAAV2-CMV-GFP-BGHpA
FEATURES             Location/Qualifiers
     source          1..4574
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   8..137
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     protein_bind    152..176
                     /gene="mutant version of attB"
                     /label=attB2
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     enhancer        253..556
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        557..760
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     711..731
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     regulatory      807..816
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             join(813..815,816..818,819..1532)
                     /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(813..815,816..818,819..1529)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(858..879)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1119..1138)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1466..1487
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     polyA_signal    1542..1766
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     protein_bind    complement(1797..1821)
                     /gene="mutant version of attB"
                     /label=attB1
                     /bound_moiety="BP Clonase(TM)"
                     /note="recombination site for the Gateway(R) BP reaction"
     repeat_region   1844..1984
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   1844..1973
                     /label=AAV2 ITR
     rep_origin      2059..2514
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2146..2165)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2356..2377
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(2531..2550)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     2650..2672
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(2710..2728)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        2796..2900
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2901..2969,2970..3761)
                     /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(3119..3138)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      3932..4520
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4421..4440
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cacatgtcct gcaggcagct gcgcgctcgc tcgctcactg aggccgcccg ggcgtcgggc
       61 gacctttggt cgcccggcct cagtgagcga gcgagcgcgc agagagggag tggccaactc
      121 catcactagg ggttcctgcg gccgcacgcg taccactttg tacaagaaag ctgggtctag
      181 atatctcgag gtcagttatt aatagtaatc aattacgggg tcattagttc atagcccata
      241 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
      301 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
      361 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
      421 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
      481 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
      541 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
      601 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
      661 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
      721 cggtaggcgt gtacggtggg aggtctatat aagcagagct ggtttagtga accgtcagat
      781 cgacggtatc gataagcttg attcgagcca ccatggtgag caagggcgag gagctgttca
      841 ccggggtggt gcccatcctg gtcgagctgg acggcgacgt aaacggccac aagttcagcg
      901 tgtccggcga gggcgagggc gatgccacct acggcaagct gaccctgaag ttcatctgca
      961 ccaccggcaa gctgcccgtg ccctggccca ccctcgtgac caccctgacc tacggcgtgc
     1021 agtgcttcag ccgctacccc gaccacatga agcagcacga cttcttcaag tccgccatgc
     1081 ccgaaggcta cgtccaggag cgcaccatct tcttcaagga cgacggcaac tacaagaccc
     1141 gcgccgaggt gaagttcgag ggcgacaccc tggtgaaccg catcgagctg aagggcatcg
     1201 acttcaagga ggacggcaac atcctggggc acaagctgga gtacaactac aacagccaca
     1261 acgtctatat catggccgac aagcagaaga acggcatcaa ggtgaacttc aagatccgcc
     1321 acaacatcga ggacggcagc gtgcagctcg ccgaccacta ccagcagaac acccccatcg
     1381 gcgacggccc cgtgctgctg cccgacaacc actacctgag cacccagtcc gccctgagca
     1441 aagaccccaa cgagaagcgc gatcacatgg tcctgctgga gttcgtgacc gccgccggga
     1501 tcactctcgg catggacgag ctgtacaagt aaatcgaatt cctgtgcctt ctagttgcca
     1561 gccatctgtt gtttgcccct cccccgtgcc ttccttgacc ctggaaggtg ccactcccac
     1621 tgtcctttcc taataaaatg aggaaattgc atcgcattgt ctgagtaggt gtcattctat
     1681 tctggggggt ggggtggggc aggacagcaa gggggaggat tgggaagaca atagcaggca
     1741 tgctggggat gcggtgggct ctatggggat ccagtcgact gaattggttc ctttaaagcc
     1801 tgcttttttg tacaaacttg tcacgtgcgg accgagcggc cgcaggaacc cctagtgatg
     1861 gagttggcca ctccctctct gcgcgctcgc tcgctcactg aggccgggcg accaaaggtc
     1921 gcccgacgcc cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg cagctgcctg
     1981 caggggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc acaccgcata
     2041 cgtcaaagca accatagtac gcgccctgta gcggcgcatt aagcgcggcg ggtgtggtgg
     2101 ttacgcgcag cgtgaccgct acacttgcca gcgccttagc gcccgctcct ttcgctttct
     2161 tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat cgggggctcc
     2221 ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt gatttgggtg
     2281 atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg acgttggagt
     2341 ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac tctatctcgg
     2401 gctattcttt tgatttataa gggattttgc cgatttcggt ctattggtta aaaaatgagc
     2461 tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgtttaca attttatggt
     2521 gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga cacccgccaa
     2581 cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac agacaagctg
     2641 tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg aaacgcgcga
     2701 gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata ataatggttt
     2761 cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     2821 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     2881 aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt attccctttt
     2941 ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa gtaaaagatg
     3001 ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac agcggtaaga
     3061 tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt aaagttctgc
     3121 tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt cgccgcatac
     3181 actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat cttacggatg
     3241 gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac actgcggcca
     3301 acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg cacaacatgg
     3361 gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc ataccaaacg
     3421 acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa ctattaactg
     3481 gcgaactact tactctagct tcccggcaac aattaataga ctggatggag gcggataaag
     3541 ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct gataaatctg
     3601 gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat ggtaagccct
     3661 cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa cgaaatagac
     3721 agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac caagtttact
     3781 catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc taggtgaaga
     3841 tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc cactgagcgt
     3901 cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg cgcgtaatct
     3961 gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg gatcaagagc
     4021 taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca aatactgttc
     4081 ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg cctacatacc
     4141 tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg tgtcttaccg
     4201 ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga acggggggtt
     4261 cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac ctacagcgtg
     4321 agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat ccggtaagcg
     4381 gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc tggtatcttt
     4441 atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga tgctcgtcag
     4501 gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc ctggcctttt
     4561 gctggccttt tgct
//
