LOCUS       sequence-418475-        5077 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  EGFP reporter vector of iTango system for AAV production.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5077)
  AUTHORS   Lee D, Creed M, Jung K, Stefanelli T, Wendler DJ, Oh WC, Mignocchi 
            NL, Luscher C, Kwon HB
  TITLE     Temporally precise labeling and control of neuromodulatory circuits 
            in the mammalian brain.
  JOURNAL   Nat Methods. 2017 May;14(5):495-503. doi: 10.1038/nmeth.4234. Epub 
            2017 Apr 3.
  PUBMED    28369042
REFERENCE   2  (bases 1 to 5077)
  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: 
            "10.1038/nmeth.4234"; journalName: "Nat Methods"; date: "2017-05"; 
            volume: "14"; issue: "5"; pages: "495-503"
COMMENT     Sequence Label: pAAV-TRE-EGFP
FEATURES             Location/Qualifiers
     source          1..5077
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     protein_bind    join(287..305,306..328,329..347,348..370,371..389,390..412,
                     413..431,432..454,455..473,474..496,497..515,516..538,
                     539..557)
                     /label=tetracycline response element
                     /bound_moiety="tetracycline repressor TetR"
                     /note="contains seven copies of the tetracycline operator 
                     tetO"
     promoter        590..628
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     CDS             join(749..751,752..754,755..1468)
                     /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(749..751,752..754,755..1465)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(794..815)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1055..1074)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1402..1423
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    1481..2069
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1534..1554)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1952..1963)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2104..2121)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2110..2317
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2340..2480
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2340..2469
                     /label=AAV2 ITR
     rep_origin      2555..3010
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2642..2661)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2852..2873
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3027..3046)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3146..3168
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3206..3224)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3292..3396
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3397..3465,3466..4257)
                     /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(3615..3634)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4428..5016
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4917..4936
                     /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 gcgaaacaat tctgcaggaa ttcggggccg cggaggctgg
      181 atcggtcccg gtgtcttcta tggaggtcaa aacagcgtgg atggcgtctc caggcgatct
      241 gacggttcac taaacgagct ctgcttatat aggtcgagtt taccactccc tatcagtgat
      301 agagaaaagt gaaagtcgag tttaccactc cctatcagtg atagagaaaa gtgaaagtcg
      361 agtttaccac tccctatcag tgatagagaa aagtgaaagt cgagtttacc actccctatc
      421 agtgatagag aaaagtgaaa gtcgagttta ccactcccta tcagtgatag agaaaagtga
      481 aagtcgagtt taccactccc tatcagtgat agagaaaagt gaaagtcgag tttaccactc
      541 cctatcagtg atagagaaaa gtgaaagtcg agctcggtac ccgggtcgag gtaggcgtgt
      601 acggtgggag gcctatataa gcagagctcg tttagtgaac cgtcagatcg cctggagacg
      661 ccatccacgc tgttttgacc tccatagaag acaccgggac cgatccagcc tccgcggccc
      721 cgaattcgag ctcggtaccc ggggatccat ggtgagcaag ggcgaggagc tgttcaccgg
      781 ggtggtgccc atcctggtcg agctggacgg cgacgtaaac ggccacaagt tcagcgtgtc
      841 cggcgagggc gagggcgatg ccacctacgg caagctgacc ctgaagttca tctgcaccac
      901 cggcaagctg cccgtgccct ggcccaccct cgtgaccacc ctgacctacg gcgtgcagtg
      961 cttcagccgc taccccgacc acatgaagca gcacgacttc ttcaagtccg ccatgcccga
     1021 aggctacgtc caggagcgca ccatcttctt caaggacgac ggcaactaca agacccgcgc
     1081 cgaggtgaag ttcgagggcg acaccctggt gaaccgcatc gagctgaagg gcatcgactt
     1141 caaggaggac ggcaacatcc tggggcacaa gctggagtac aactacaaca gccacaacgt
     1201 ctatatcatg gccgacaagc agaagaacgg catcaaggtg aacttcaaga tccgccacaa
     1261 catcgaggac ggcagcgtgc agctcgccga ccactaccag cagaacaccc ccatcggcga
     1321 cggccccgtg ctgctgcccg acaaccacta cctgagcacc cagtccgccc tgagcaaaga
     1381 ccccaacgag aagcgcgatc acatggtcct gctggagttc gtgaccgccg ccgggatcac
     1441 tctcggcatg gacgagctgt acaagtagaa gcttatcgat aatcaacctc tggattacaa
     1501 aatttgtgaa agattgactg gtattcttaa ctatgttgct ccttttacgc tatgtggata
     1561 cgctgcttta atgcctttgt atcatgctat tgcttcccgt atggctttca ttttctcctc
     1621 cttgtataaa tcctggttgc tgtctcttta tgaggagttg tggcccgttg tcaggcaacg
     1681 tggcgtggtg tgcactgtgt ttgctgacgc aacccccact ggttggggca ttgccaccac
     1741 ctgtcagctc ctttccggga ctttcgcttt ccccctccct attgccacgg cggaactcat
     1801 cgccgcctgc cttgcccgct gctggacagg ggctcggctg ttgggcactg acaattccgt
     1861 ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc gcctatgttg ccacctggat
     1921 tctgcgcggg acgtccttct gctacgtccc ttcggccctc aatccagcgg accttccttc
     1981 ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag
     2041 tcggatctcc ctttgggccg cctccccgca tcgataccga gcgctgctcg agagatcgat
     2101 ctgcctcgac tgtgccttct agttgccagc catctgttgt ttgcccctcc cccgtgcctt
     2161 ccttgaccct ggaaggtgcc actcccactg tcctttccta ataaaatgag gaaattgcat
     2221 cgcattgtct gagtaggtgt cattctattc tggggggtgg ggtggggcag gacagcaagg
     2281 gggaggattg ggaagacaat agcaggcatg ctggggacac gtgcggaccg agcggccgca
     2341 ggaaccccta gtgatggagt tggccactcc ctctctgcgc gctcgctcgc tcactgaggc
     2401 cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag tgagcgagcg
     2461 agcgcgcagc tgcctgcagg ggcgcctgat gcggtatttt ctccttacgc atctgtgcgg
     2521 tatttcacac cgcatacgtc aaagcaacca tagtacgcgc cctgtagcgg cgcattaagc
     2581 gcggcgggtg tggtggttac gcgcagcgtg accgctacac ttgccagcgc cttagcgccc
     2641 gctcctttcg ctttcttccc ttcctttctc gccacgttcg ccggctttcc ccgtcaagct
     2701 ctaaatcggg ggctcccttt agggttccga tttagtgctt tacggcacct cgaccccaaa
     2761 aaacttgatt tgggtgatgg ttcacgtagt gggccatcgc cctgatagac ggtttttcgc
     2821 cctttgacgt tggagtccac gttctttaat agtggactct tgttccaaac tggaacaaca
     2881 ctcaactcta tctcgggcta ttcttttgat ttataaggga ttttgccgat ttcggtctat
     2941 tggttaaaaa atgagctgat ttaacaaaaa tttaacgcga attttaacaa aatattaacg
     3001 tttacaattt tatggtgcac tctcagtaca atctgctctg atgccgcata gttaagccag
     3061 ccccgacacc cgccaacacc cgctgacgcg ccctgacggg cttgtctgct cccggcatcc
     3121 gcttacagac aagctgtgac cgtctccggg agctgcatgt gtcagaggtt ttcaccgtca
     3181 tcaccgaaac gcgcgagacg aaagggcctc gtgatacgcc tatttttata ggttaatgtc
     3241 atgataataa tggtttctta gacgtcaggt ggcacttttc ggggaaatgt gcgcggaacc
     3301 cctatttgtt tatttttcta aatacattca aatatgtatc cgctcatgag acaataaccc
     3361 tgataaatgc ttcaataata ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc
     3421 gcccttattc ccttttttgc ggcattttgc cttcctgttt ttgctcaccc agaaacgctg
     3481 gtgaaagtaa aagatgctga agatcagttg ggtgcacgag tgggttacat cgaactggat
     3541 ctcaacagcg gtaagatcct tgagagtttt cgccccgaag aacgttttcc aatgatgagc
     3601 acttttaaag ttctgctatg tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa
     3661 ctcggtcgcc gcatacacta ttctcagaat gacttggttg agtactcacc agtcacagaa
     3721 aagcatctta cggatggcat gacagtaaga gaattatgca gtgctgccat aaccatgagt
     3781 gataacactg cggccaactt acttctgaca acgatcggag gaccgaagga gctaaccgct
     3841 tttttgcaca acatggggga tcatgtaact cgccttgatc gttgggaacc ggagctgaat
     3901 gaagccatac caaacgacga gcgtgacacc acgatgcctg tagcaatggc aacaacgttg
     3961 cgcaaactat taactggcga actacttact ctagcttccc ggcaacaatt aatagactgg
     4021 atggaggcgg ataaagttgc aggaccactt ctgcgctcgg cccttccggc tggctggttt
     4081 attgctgata aatctggagc cggtgagcgt gggtctcgcg gtatcattgc agcactgggg
     4141 ccagatggta agccctcccg tatcgtagtt atctacacga cggggagtca ggcaactatg
     4201 gatgaacgaa atagacagat cgctgagata ggtgcctcac tgattaagca ttggtaactg
     4261 tcagaccaag tttactcata tatactttag attgatttaa aacttcattt ttaatttaaa
     4321 aggatctagg tgaagatcct ttttgataat ctcatgacca aaatccctta acgtgagttt
     4381 tcgttccact gagcgtcaga ccccgtagaa aagatcaaag gatcttcttg agatcctttt
     4441 tttctgcgcg taatctgctg cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt
     4501 ttgccggatc aagagctacc aactcttttt ccgaaggtaa ctggcttcag cagagcgcag
     4561 ataccaaata ctgttcttct agtgtagccg tagttaggcc accacttcaa gaactctgta
     4621 gcaccgccta catacctcgc tctgctaatc ctgttaccag tggctgctgc cagtggcgat
     4681 aagtcgtgtc ttaccgggtt ggactcaaga cgatagttac cggataaggc gcagcggtcg
     4741 ggctgaacgg ggggttcgtg cacacagccc agcttggagc gaacgaccta caccgaactg
     4801 agatacctac agcgtgagct atgagaaagc gccacgcttc ccgaagggag aaaggcggac
     4861 aggtatccgg taagcggcag ggtcggaaca ggagagcgca cgagggagct tccaggggga
     4921 aacgcctggt atctttatag tcctgtcggg tttcgccacc tctgacttga gcgtcgattt
     4981 ttgtgatgct cgtcaggggg gcggagccta tggaaaaacg ccagcaacgc ggccttttta
     5041 cggttcctgg ccttttgctg gccttttgct cacatgt
//
