LOCUS       sequence-177102-        5088 bp    DNA     circular SYN 05-MAY-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 5088)
  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 5088)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 5, 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..5088
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     protein_bind    join(298..316,317..339,340..358,359..381,382..400,401..423,
                     424..442,443..465,466..484,485..507,508..526,527..549,
                     550..568)
                     /label=tetracycline response element
                     /bound_moiety="tetracycline repressor TetR"
                     /note="contains seven copies of the tetracycline operator 
                     tetO"
     promoter        601..639
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     CDS             join(760..762,763..765,766..1479)
                     /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(760..762,763..765,766..1476)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(805..826)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1066..1085)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1413..1434
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    1492..2080
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1545..1565)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(1963..1974)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(2115..2132)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2121..2328
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   2351..2491
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2351..2480
                     /label=AAV2 ITR
     rep_origin      2566..3021
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2653..2672)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     2863..2884
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3038..3057)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3157..3179
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3217..3235)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3303..3407
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3408..3476,3477..4268)
                     /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(3626..3645)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4439..5027
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4928..4947
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgaaacaa ttctgcagga attcggggcc
      181 gcggaggctg gatcggtccc ggtgtcttct atggaggtca aaacagcgtg gatggcgtct
      241 ccaggcgatc tgacggttca ctaaacgagc tctgcttata taggtcgagt ttaccactcc
      301 ctatcagtga tagagaaaag tgaaagtcga gtttaccact ccctatcagt gatagagaaa
      361 agtgaaagtc gagtttacca ctccctatca gtgatagaga aaagtgaaag tcgagtttac
      421 cactccctat cagtgataga gaaaagtgaa agtcgagttt accactccct atcagtgata
      481 gagaaaagtg aaagtcgagt ttaccactcc ctatcagtga tagagaaaag tgaaagtcga
      541 gtttaccact ccctatcagt gatagagaaa agtgaaagtc gagctcggta cccgggtcga
      601 ggtaggcgtg tacggtggga ggcctatata agcagagctc gtttagtgaa ccgtcagatc
      661 gcctggagac gccatccacg ctgttttgac ctccatagaa gacaccggga ccgatccagc
      721 ctccgcggcc ccgaattcga gctcggtacc cggggatcca tggtgagcaa gggcgaggag
      781 ctgttcaccg gggtggtgcc catcctggtc gagctggacg gcgacgtaaa cggccacaag
      841 ttcagcgtgt ccggcgaggg cgagggcgat gccacctacg gcaagctgac cctgaagttc
      901 atctgcacca ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cctgacctac
      961 ggcgtgcagt gcttcagccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc
     1021 gccatgcccg aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac
     1081 aagacccgcg ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag
     1141 ggcatcgact tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac
     1201 agccacaacg tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag
     1261 atccgccaca acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc
     1321 cccatcggcg acggccccgt gctgctgccc gacaaccact acctgagcac ccagtccgcc
     1381 ctgagcaaag accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc
     1441 gccgggatca ctctcggcat ggacgagctg tacaagtaga agcttatcga taatcaacct
     1501 ctggattaca aaatttgtga aagattgact ggtattctta actatgttgc tccttttacg
     1561 ctatgtggat acgctgcttt aatgcctttg tatcatgcta ttgcttcccg tatggctttc
     1621 attttctcct ccttgtataa atcctggttg ctgtctcttt atgaggagtt gtggcccgtt
     1681 gtcaggcaac gtggcgtggt gtgcactgtg tttgctgacg caacccccac tggttggggc
     1741 attgccacca cctgtcagct cctttccggg actttcgctt tccccctccc tattgccacg
     1801 gcggaactca tcgccgcctg ccttgcccgc tgctggacag gggctcggct gttgggcact
     1861 gacaattccg tggtgttgtc ggggaaatca tcgtcctttc cttggctgct cgcctatgtt
     1921 gccacctgga ttctgcgcgg gacgtccttc tgctacgtcc cttcggccct caatccagcg
     1981 gaccttcctt cccgcggcct gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc
     2041 cctcagacga gtcggatctc cctttgggcc gcctccccgc atcgataccg agcgctgctc
     2101 gagagatcga tctgcctcga ctgtgccttc tagttgccag ccatctgttg tttgcccctc
     2161 ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct aataaaatga
     2221 ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg gggtggggca
     2281 ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggaca cgtgcggacc
     2341 gagcggccgc aggaacccct agtgatggag ttggccactc cctctctgcg cgctcgctcg
     2401 ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg ggcggcctca
     2461 gtgagcgagc gagcgcgcag ctgcctgcag gggcgcctga tgcggtattt tctccttacg
     2521 catctgtgcg gtatttcaca ccgcatacgt caaagcaacc atagtacgcg ccctgtagcg
     2581 gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt gaccgctaca cttgccagcg
     2641 ccttagcgcc cgctcctttc gctttcttcc cttcctttct cgccacgttc gccggctttc
     2701 cccgtcaagc tctaaatcgg gggctccctt tagggttccg atttagtgct ttacggcacc
     2761 tcgaccccaa aaaacttgat ttgggtgatg gttcacgtag tgggccatcg ccctgataga
     2821 cggtttttcg ccctttgacg ttggagtcca cgttctttaa tagtggactc ttgttccaaa
     2881 ctggaacaac actcaactct atctcgggct attcttttga tttataaggg attttgccga
     2941 tttcggtcta ttggttaaaa aatgagctga tttaacaaaa atttaacgcg aattttaaca
     3001 aaatattaac gtttacaatt ttatggtgca ctctcagtac aatctgctct gatgccgcat
     3061 agttaagcca gccccgacac ccgccaacac ccgctgacgc gccctgacgg gcttgtctgc
     3121 tcccggcatc cgcttacaga caagctgtga ccgtctccgg gagctgcatg tgtcagaggt
     3181 tttcaccgtc atcaccgaaa cgcgcgagac gaaagggcct cgtgatacgc ctatttttat
     3241 aggttaatgt catgataata atggtttctt agacgtcagg tggcactttt cggggaaatg
     3301 tgcgcggaac ccctatttgt ttatttttct aaatacattc aaatatgtat ccgctcatga
     3361 gacaataacc ctgataaatg cttcaataat attgaaaaag gaagagtatg agtattcaac
     3421 atttccgtgt cgcccttatt cccttttttg cggcattttg ccttcctgtt tttgctcacc
     3481 cagaaacgct ggtgaaagta aaagatgctg aagatcagtt gggtgcacga gtgggttaca
     3541 tcgaactgga tctcaacagc ggtaagatcc ttgagagttt tcgccccgaa gaacgttttc
     3601 caatgatgag cacttttaaa gttctgctat gtggcgcggt attatcccgt attgacgccg
     3661 ggcaagagca actcggtcgc cgcatacact attctcagaa tgacttggtt gagtactcac
     3721 cagtcacaga aaagcatctt acggatggca tgacagtaag agaattatgc agtgctgcca
     3781 taaccatgag tgataacact gcggccaact tacttctgac aacgatcgga ggaccgaagg
     3841 agctaaccgc ttttttgcac aacatggggg atcatgtaac tcgccttgat cgttgggaac
     3901 cggagctgaa tgaagccata ccaaacgacg agcgtgacac cacgatgcct gtagcaatgg
     3961 caacaacgtt gcgcaaacta ttaactggcg aactacttac tctagcttcc cggcaacaat
     4021 taatagactg gatggaggcg gataaagttg caggaccact tctgcgctcg gcccttccgg
     4081 ctggctggtt tattgctgat aaatctggag ccggtgagcg tgggtctcgc ggtatcattg
     4141 cagcactggg gccagatggt aagccctccc gtatcgtagt tatctacacg acggggagtc
     4201 aggcaactat ggatgaacga aatagacaga tcgctgagat aggtgcctca ctgattaagc
     4261 attggtaact gtcagaccaa gtttactcat atatacttta gattgattta aaacttcatt
     4321 tttaatttaa aaggatctag gtgaagatcc tttttgataa tctcatgacc aaaatccctt
     4381 aacgtgagtt ttcgttccac tgagcgtcag accccgtaga aaagatcaaa ggatcttctt
     4441 gagatccttt ttttctgcgc gtaatctgct gcttgcaaac aaaaaaacca ccgctaccag
     4501 cggtggtttg tttgccggat caagagctac caactctttt tccgaaggta actggcttca
     4561 gcagagcgca gataccaaat actgttcttc tagtgtagcc gtagttaggc caccacttca
     4621 agaactctgt agcaccgcct acatacctcg ctctgctaat cctgttacca gtggctgctg
     4681 ccagtggcga taagtcgtgt cttaccgggt tggactcaag acgatagtta ccggataagg
     4741 cgcagcggtc gggctgaacg gggggttcgt gcacacagcc cagcttggag cgaacgacct
     4801 acaccgaact gagataccta cagcgtgagc tatgagaaag cgccacgctt cccgaaggga
     4861 gaaaggcgga caggtatccg gtaagcggca gggtcggaac aggagagcgc acgagggagc
     4921 ttccaggggg aaacgcctgg tatctttata gtcctgtcgg gtttcgccac ctctgacttg
     4981 agcgtcgatt tttgtgatgc tcgtcagggg ggcggagcct atggaaaaac gccagcaacg
     5041 cggccttttt acggttcctg gccttttgct ggccttttgc tcacatgt
//
