LOCUS       sequence-485694-        6325 bp    DNA     circular SYN 10-SEP-2025
DEFINITION  AAV version of Tet inducible control plasmid.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6325)
  AUTHORS   Bareghamyan A, Deng C, Daoudi S, Yadav SC, Lu X, Zhang W, Campbell 
            RE, Kramer RH, Chenoweth DM, Arnold DB
  TITLE     A toolbox for ablating excitatory and inhibitory synapses.
  JOURNAL   eLife. 2025 Apr 29;13:RP103757. doi: 10.7554/eLife.103757.
  PUBMED    40298577
REFERENCE   2  (bases 1 to 6325)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 10, 2025 from SnapGene Server 8.0.1
            https://www.snapgene.com
COMMENT     SGRef: number: 1; type: "Journal Article"; journalName: "eLife."; 
            date: "2025-04-29"; volume: "13:RP103757. doi"; pages: " 
            10.7554/eLife.103757"
COMMENT     Sequence Label: AAV-TRE-RandE3-P2A-GFP
FEATURES             Location/Qualifiers
     source          1..6325
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        148..462
                     /label=tight TRE promoter
                     /note="Tet-responsive promoter PTight, consisting of seven 
                     tet operator sequences followed by the minimal CMV 
                     promoter"
     protein_bind    156..174
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    192..210
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    227..245
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    263..281
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    299..317
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    334..352
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    370..388
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     435..459
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             755..781
                     /codon_start=1
                     /product="HA (human influenza hemagglutinin) epitope tag"
                     /label=HA
                     /translation="YPYDVPDYA"
     CDS             1673..1729
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             join(1742..1744,1745..2455)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDEL*K"
     primer_bind     complement(1784..1805)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2045..2064)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2392..2413
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     misc_feature    2465..3053
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2518..2538)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2936..2947)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3072..3548
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3271..3290)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3588..3728
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3588..3717
                     /label=AAV2 ITR
     rep_origin      3803..4258
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3890..3909)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4100..4121
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4275..4294)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4394..4416
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4454..4472)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4540..4644
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4645..4713,4714..5505)
                     /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(4863..4882)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5676..6264
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6165..6184
                     /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 gcgtctcgag tttactccct atcagtgata gagaacgtat
      181 gtcgagttta ctccctatca gtgatagaga acgatgtcga gtttactccc tatcagtgat
      241 agagaacgta tgtcgagttt actccctatc agtgatagag aacgtatgtc gagtttactc
      301 cctatcagtg atagagaacg tatgtcgagt ttatccctat cagtgataga gaacgtatgt
      361 cgagtttact ccctatcagt gatagagaac gtatgtcgag gtaggcgtgt acggtgggag
      421 gcctatataa gcagagctcg tttagtgaac cgtcagatcg cctggagaat tctctagaat
      481 gctcgaagtc gtcgttgcga ctccgaccag cctcctaatc agctggcggg cgacgcagga
      541 gcacgcgcgc tactaccgca tcacctacgg tgaaactggt ggcaatagcc ctgtccagga
      601 attcaccgtg cctggcagca agtccactgc taccatcagc ggcctgaaac ctggtgtcga
      661 ctataccatc acggtgtacg ccgtcacgcg cgagccccag gccagctccg accagtcccc
      721 gatctccatc aactaccgca ccagatcaac cggttaccca tacgatgttc cagattacgc
      781 tggtggaagc acgcgttgtg tgatttgcca ggggcggcct aaaaatggtt gcattgttca
      841 cggcaaaacc ggacacctga tgtcatgttt cacgtgtgca aaggcactag ctgcggcgaa
      901 tgcgccctgc cccgtgtgca gagctagcaa gctctcaggt ggcggaggga gcggaggagg
      961 tggctctgga ggtggtggct cagctgaagc atggtacaat ctgggaaacg cttattacaa
     1021 gcagggcgac tatcaaaaag ccattgagta ttaccaaaag gctcttgagc tagaccctaa
     1081 taacgccgaa gcgtggtata acctaggcaa cgcatattac aagcagggag actaccaaaa
     1141 ggcaatcgaa tattaccaga aggcactcga actggaccca aataacgcag aggcttggta
     1201 taatctggga aacgcctact ataaacaggg tgactaccaa aaggcgatag aggactatca
     1261 gaaggccttg gagctcgatc cgaacaatct gcaacgtagt gctggtggcg gagggtctgg
     1321 tggaggaggc tcaggaggag gtggcgcatc aggtaccaag aagaaactca gcaagtcaga
     1381 catcatgttg gtgcagagcg ccaatgtccc tagcaacccg acccaggtgc cggtagagga
     1441 gtccggaggc ttcggttccc atcaccacaa tcagaactat tgctatcagg tctgcctcac
     1501 cccagagtcc gccaagaccg acctgatgtt cttgaagccc tgcagccctt cccggagtac
     1561 agacgctgag cacaatccct gtggagccat cgtcaccggg tacagcgacc agcaaccaga
     1621 catcatttcc aacggaagca ttttgtccaa cgagaatggt ggatcatgta cagctactaa
     1681 cttcagcctg ctgaagcagg ctggagacgt ggaggagaac cctggacctg gaggtactgg
     1741 ggtgagcaag ggcgaggagc tgttcaccgg ggtggtgccc atcctggtcg agctggacgg
     1801 cgacgtaaac ggccacaagt tcagcgtgtc cggcgagggc gagggcgatg ccacctacgg
     1861 caagctgacc ctgaagttca tctgcaccac cggcaagctg cccgtgccct ggcccaccct
     1921 cgtgaccacc ctgacctacg gcgtgcagtg cttcagccgc taccccgacc acatgaagca
     1981 gcacgacttc ttcaagtccg ccatgcccga aggctacgtc caggagcgca ccatcttctt
     2041 caaggacgac ggcaactaca agacccgcgc cgaggtgaag ttcgagggcg acaccctggt
     2101 gaaccgcatc gagctgaagg gcatcgactt caaggaggac ggcaacatcc tggggcacaa
     2161 gctggagtac aactacaaca gccacaacgt ctatatcatg gccgacaagc agaagaacgg
     2221 catcaaggtg aacttcaaga tccgccacaa catcgaggac ggcagcgtgc agctcgccga
     2281 ccactaccag cagaacaccc ccatcggcga cggccccgtg ctgctgcccg acaaccacta
     2341 cctgagcacc cagtccgccc tgagcaaaga ccccaacgag aagcgcgatc acatggtcct
     2401 gctggagttc gtgaccgccg ccgggatcac tctcggcatg gacgagctgt aaaagcttat
     2461 cgataatcaa cctctggatt acaaaatttg tgaaagattg actggtattc ttaactatgt
     2521 tgctcctttt acgctatgtg gatacgctgc tttaatgcct ttgtatcatg ctattgcttc
     2581 ccgtatggct ttcattttct cctccttgta taaatcctgg ttgctgtctc tttatgagga
     2641 gttgtggccc gttgtcaggc aacgtggcgt ggtgtgcact gtgtttgctg acgcaacccc
     2701 cactggttgg ggcattgcca ccacctgtca gctcctttcc gggactttcg ctttccccct
     2761 ccctattgcc acggcggaac tcatcgccgc ctgccttgcc cgctgctgga caggggctcg
     2821 gctgttgggc actgacaatt ccgtggtgtt gtcggggaaa tcatcgtcct ttccttggct
     2881 gctcgcctat gttgccacct ggattctgcg cgggacgtcc ttctgctacg tcccttcggc
     2941 cctcaatcca gcggaccttc cttcccgcgg cctgctgccg gctctgcggc ctcttccgcg
     3001 tcttcgcctt cgccctcaga cgagtcggat ctccctttgg gccgcctccc cgcatcgata
     3061 ccgagatcta cgggtggcat ccctgtgacc cctccccagt gcctctcctg gccctggaag
     3121 ttgccactcc agtgcccacc agccttgtcc taataaaatt aagttgcatc attttgtctg
     3181 actaggtgtc cttctataat attatggggt ggaggggggt ggtatggagc aaggggcaag
     3241 ttgggaagac aacctgtagg gcctgcgggg tctattggga accaagctgg agtgcagtgg
     3301 cacaatcttg gctcactgca atctccgcct cctgggttca agcgattctc ctgcctcagc
     3361 ctcccgagtt gttgggattc caggcatgca tgaccaggct cagctaattt ttgttttttt
     3421 ggtagagacg gggtttcacc atattggcca ggctggtctc caactcctaa tctcaggtga
     3481 tctacccacc ttggcctccc aaattgctgg gattacaggc gtgaaccact gctcccttcc
     3541 ctgtccttct gattttgtag gtaaccacgt gcggaccgag cggccgcagg aacccctagt
     3601 gatggagttg gccactccct ctctgcgcgc tcgctcgctc actgaggccg ggcgaccaaa
     3661 ggtcgcccga cgcccgggct ttgcccgggc ggcctcagtg agcgagcgag cgcgcagctg
     3721 cctgcagggg cgcctgatgc ggtattttct ccttacgcat ctgtgcggta tttcacaccg
     3781 catacgtcaa agcaaccata gtacgcgccc tgtagcggcg cattaagcgc ggcgggtgtg
     3841 gtggttacgc gcagcgtgac cgctacactt gccagcgcct tagcgcccgc tcctttcgct
     3901 ttcttccctt cctttctcgc cacgttcgcc ggctttcccc gtcaagctct aaatcggggg
     3961 ctccctttag ggttccgatt tagtgcttta cggcacctcg accccaaaaa acttgatttg
     4021 ggtgatggtt cacgtagtgg gccatcgccc tgatagacgg tttttcgccc tttgacgttg
     4081 gagtccacgt tctttaatag tggactcttg ttccaaactg gaacaacact caactctatc
     4141 tcgggctatt cttttgattt ataagggatt ttgccgattt cggtctattg gttaaaaaat
     4201 gagctgattt aacaaaaatt taacgcgaat tttaacaaaa tattaacgtt tacaatttta
     4261 tggtgcactc tcagtacaat ctgctctgat gccgcatagt taagccagcc ccgacacccg
     4321 ccaacacccg ctgacgcgcc ctgacgggct tgtctgctcc cggcatccgc ttacagacaa
     4381 gctgtgaccg tctccgggag ctgcatgtgt cagaggtttt caccgtcatc accgaaacgc
     4441 gcgagacgaa agggcctcgt gatacgccta tttttatagg ttaatgtcat gataataatg
     4501 gtttcttaga cgtcaggtgg cacttttcgg ggaaatgtgc gcggaacccc tatttgttta
     4561 tttttctaaa tacattcaaa tatgtatccg ctcatgagac aataaccctg ataaatgctt
     4621 caataatatt gaaaaaggaa gagtatgagt attcaacatt tccgtgtcgc ccttattccc
     4681 ttttttgcgg cattttgcct tcctgttttt gctcacccag aaacgctggt gaaagtaaaa
     4741 gatgctgaag atcagttggg tgcacgagtg ggttacatcg aactggatct caacagcggt
     4801 aagatccttg agagttttcg ccccgaagaa cgttttccaa tgatgagcac ttttaaagtt
     4861 ctgctatgtg gcgcggtatt atcccgtatt gacgccgggc aagagcaact cggtcgccgc
     4921 atacactatt ctcagaatga cttggttgag tactcaccag tcacagaaaa gcatcttacg
     4981 gatggcatga cagtaagaga attatgcagt gctgccataa ccatgagtga taacactgcg
     5041 gccaacttac ttctgacaac gatcggagga ccgaaggagc taaccgcttt tttgcacaac
     5101 atgggggatc atgtaactcg ccttgatcgt tgggaaccgg agctgaatga agccatacca
     5161 aacgacgagc gtgacaccac gatgcctgta gcaatggcaa caacgttgcg caaactatta
     5221 actggcgaac tacttactct agcttcccgg caacaattaa tagactggat ggaggcggat
     5281 aaagttgcag gaccacttct gcgctcggcc cttccggctg gctggtttat tgctgataaa
     5341 tctggagccg gtgagcgtgg gtctcgcggt atcattgcag cactggggcc agatggtaag
     5401 ccctcccgta tcgtagttat ctacacgacg gggagtcagg caactatgga tgaacgaaat
     5461 agacagatcg ctgagatagg tgcctcactg attaagcatt ggtaactgtc agaccaagtt
     5521 tactcatata tactttagat tgatttaaaa cttcattttt aatttaaaag gatctaggtg
     5581 aagatccttt ttgataatct catgaccaaa atcccttaac gtgagttttc gttccactga
     5641 gcgtcagacc ccgtagaaaa gatcaaagga tcttcttgag atcctttttt tctgcgcgta
     5701 atctgctgct tgcaaacaaa aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa
     5761 gagctaccaa ctctttttcc gaaggtaact ggcttcagca gagcgcagat accaaatact
     5821 gttcttctag tgtagccgta gttaggccac cacttcaaga actctgtagc accgcctaca
     5881 tacctcgctc tgctaatcct gttaccagtg gctgctgcca gtggcgataa gtcgtgtctt
     5941 accgggttgg actcaagacg atagttaccg gataaggcgc agcggtcggg ctgaacgggg
     6001 ggttcgtgca cacagcccag cttggagcga acgacctaca ccgaactgag atacctacag
     6061 cgtgagctat gagaaagcgc cacgcttccc gaagggagaa aggcggacag gtatccggta
     6121 agcggcaggg tcggaacagg agagcgcacg agggagcttc cagggggaaa cgcctggtat
     6181 ctttatagtc ctgtcgggtt tcgccacctc tgacttgagc gtcgattttt gtgatgctcg
     6241 tcaggggggc ggagcctatg gaaaaacgcc agcaacgcgg cctttttacg gttcctggcc
     6301 ttttgctggc cttttgctca catgt
//
