LOCUS       sequence-486961-        6901 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bidirectional AAV transfer plasmid expressing hSyn-tTA and 
            TRE-FLEX-mNeonGreen-TY1 in the other direction..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6901)
  AUTHORS   E11 Bio
  TITLE     E11 Bio PRISM Plasmids
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 6901)
  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: 
            "Unpublished"
COMMENT     Sequence Label: pAAV-BD-hSynTet-TRE-DIO-mNeonGreen-TY1
FEATURES             Location/Qualifiers
     source          1..6901
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    948..966
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    985..1003
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1022..1040
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1059..1077
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1096..1114
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1133..1151
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1170..1188
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    1207..1225
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     1247..1271
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    complement(1370..1403)
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    complement(1454..1487)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      2070..2079
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2331..2340
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    2347..2380
                     /label=lox2272
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (AAGTATCC) (Shaw et al., 2021). lox2272 sites are 
                     compatible with each other, but incompatible with loxP or 
                     loxN sites (Lee and Saito, 1988)."
     protein_bind    2431..2464
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    2489..3077
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(2542..2562)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2960..2971)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3109..3585
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3308..3327)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3625..3765
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3625..3754
                     /label=AAV2 ITR
     rep_origin      3840..4295
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3927..3946)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4137..4158
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4312..4331)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     4431..4453
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(4491..4509)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        4577..4681
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4682..4750,4751..5542)
                     /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(4900..4919)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5713..6301
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6202..6221
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   6363..6492
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     polyA_signal    6502..6623
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(6539..6558)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     6593..6612
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(6647..6655,6656..6691,6692..6694,6695..6730,
                     6731..6733,6734..6769,6770..6775,6776..6901,1..492))
                     /codon_start=1
                     /product="improved tetracycline-controlled transactivator"
                     /label=tTA-Advanced
                     /note="In the Tet-Off(R) system, tTA-Advanced binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the absence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINSALELLNEVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALAIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEDQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDHQGAEPAFLFGLELIICGLEKQLKCESGGPADALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
ORIGIN
        1 atcggtggta ggtgtctctc tttcctcttt tgctacttga tgctcctgat cctccaatac
       61 gcagcccagt gtaaagtggc ccacggcgga cagagcgtac agtgcgttct ccagggagaa
      121 gccttgctga cacaggaacg cgagctgatt ttccagggtt tcgtactgtt tctctgttgg
      181 gcgggtgccg agatgcactt tagccccgtc gcgatgtgag aggagagcac agcggaatga
      241 cttggcgttg ttccgcagaa agtcttgcca tgactcgcct tccagggggc agaagtgggt
      301 atgatgcctg tccagcatct cgattgccag ggcatcgagc agggcccgct tgttcttcac
      361 gtgccagtac agggtaggct gctcaactcc cagcttttga gcgagtttcc ttgtcgtcag
      421 gccttcgata ccgacttcat tgagtaattc cagagcagag tttatgactt tgctcttgtc
      481 cagtctagac atggtgaatt cgatatcaag cttatcgata ccgtccgccg cagcgcagat
      541 ggtcgcgccc gtgcccccct atctcgcgcc tcgcgtggtg cggtccggct gggccggcgg
      601 cggcgcggac gcgaccaagg tggccgggaa ggggagtttg cgggggaccg gcgagtgacg
      661 tcagcgcgcc ttcagtgctg aggcggcggt ggcgcgcgcc gccaggcggg ggcgaaggca
      721 ctgtccgcgg tgctgaagct ggcagtgcgc acgcgcctcg ccgcatcctg tttcccctcc
      781 ccctctctga taggggatgc gcaatttggg gaatgggggt tgggtgcttg tccagtgggt
      841 cggggtcggt cgtcaggtag gcacccccac cccgcctcat cctggtccta aaacccactt
      901 gcactcatac gcagggccct ctgcagtatc aagtgccacc tgacgtctcc ctatcagtga
      961 tagagaagtc gacacgtctc gagctcccta tcagtgatag agaaggtacg tctagaacgt
     1021 ctccctatca gtgatagaga agtcgacacg tctcgagctc cctatcagtg atagagaagg
     1081 tacgtctaga acgtctccct atcagtgata gagaagtcga cacgtctcga gctccctatc
     1141 agtgatagag aaggtacgtc tagaacgtct ccctatcagt gatagagaag tcgacacgtc
     1201 tcgagctccc tatcagtgat agagaagcta ccccctatat aagcagagct cgtttagtga
     1261 accgtcagat cgcctggaga cgccatccac gctgttttga cctccataga agacaccggg
     1321 accgatccag cctggatccg gtaccggatc ctctagagtc gactccggaa taacttcgta
     1381 taggatactt tatacgaagt tatgcagaat ggtagctgga ttgtagctgc tattagcaat
     1441 atgaaacctc ttaataactt cgtatagcat acattatacg aagttatggc gcgcctcact
     1501 tgtccaagga attcttactt tgattatcga taagcttgac ctggctggtt tagcaacaga
     1561 actaatcgca ccagtcctac tgaagaattc ttagtccaga gggtcctggt tggtgtgcac
     1621 ctcgctgccg cccagctcgt ccatgcccat cacatcggta aaggcctttt gccactcctt
     1681 gaagttgagc tcggtcttgg agtgcttgag ctccgtctta cggaacacgt acatcggctg
     1741 gttcttcaga tagttagccg ccattggctt ggcaaaggtg taggtggtcc gcgcagtgct
     1801 ccggtagcgc ttgccatttc cagtggtgta actccactta aaggtactga tgatggtttt
     1861 gtcgttgggg taagtcttct tcgacctgca ccagtccgca gcggtcagcg agttggtcat
     1921 cacaggaccg tcagcaggga aaccagtccc cttcacctgg gcctctcctt tgatgtggct
     1981 tccctcgtag gtgtagcggt agttaacagt aagggaggca ccatcttcaa actgcattgt
     2041 gcgatggact tggtatccgg agccatctac catggcggcc tggaaaggcg acatcccgtc
     2101 agggtagggc aggtactgat ggaagccata cccgatatga gggaccagaa tccaggggga
     2161 gaactggagg tcacccttgg tggacttcag gtttaactcc tcataaccat catttggatt
     2221 gccggtgccc tgacccacca tgtcaaagtc cacaccgttg atggagccaa agatgtgtaa
     2281 ctcatgtgtc gctgggagag aggccatgtt atcctcctcg cccttgctca ccatggtggc
     2341 gctagcataa cttcgtataa agtatcctat acgaagttat ttgccttaac ccagaaatta
     2401 tcactgttat tctttagaat ggtgcaaaga ataacttcgt ataatgtatg ctatacgaag
     2461 ttatgaattc gatatcaagc ttatcgataa tcaacctctg gattacaaaa tttgtgaaag
     2521 attgactggt attcttaact atgttgctcc ttttacgcta tgtggatacg ctgctttaat
     2581 gcctttgtat catgctattg cttcccgtat ggctttcatt ttctcctcct tgtataaatc
     2641 ctggttgctg tctctttatg aggagttgtg gcccgttgtc aggcaacgtg gcgtggtgtg
     2701 cactgtgttt gctgacgcaa cccccactgg ttggggcatt gccaccacct gtcagctcct
     2761 ttccgggact ttcgctttcc ccctccctat tgccacggcg gaactcatcg ccgcctgcct
     2821 tgcccgctgc tggacagggg ctcggctgtt gggcactgac aattccgtgg tgttgtcggg
     2881 gaaatcatcg tcctttcctt ggctgctcgc ctatgttgcc acctggattc tgcgcgggac
     2941 gtccttctgc tacgtccctt cggccctcaa tccagcggac cttccttccc gcggcctgct
     3001 gccggctctg cggcctcttc cgcgtcttcg ccttcgccct cagacgagtc ggatctccct
     3061 ttgggccgcc tccccgcatc gataccgagc gctgctcgag agatctacgg gtggcatccc
     3121 tgtgacccct ccccagtgcc tctcctggcc ctggaagttg ccactccagt gcccaccagc
     3181 cttgtcctaa taaaattaag ttgcatcatt ttgtctgact aggtgtcctt ctataatatt
     3241 atggggtgga ggggggtggt atggagcaag gggcaagttg ggaagacaac ctgtagggcc
     3301 tgcggggtct attgggaacc aagctggagt gcagtggcac aatcttggct cactgcaatc
     3361 tccgcctcct gggttcaagc gattctcctg cctcagcctc ccgagttgtt gggattccag
     3421 gcatgcatga ccaggctcag ctaatttttg tttttttggt agagacgggg tttcaccata
     3481 ttggccaggc tggtctccaa ctcctaatct caggtgatct acccaccttg gcctcccaaa
     3541 ttgctgggat tacaggcgtg aaccactgct cccttccctg tccttctgat tttgtaggta
     3601 accacgtgcg gaccgagcgg ccgcaggaac ccctagtgat ggagttggcc actccctctc
     3661 tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt cgcccgacgc ccgggctttg
     3721 cccgggcggc ctcagtgagc gagcgagcgc gcagctgcct gcaggggcgc ctgatgcggt
     3781 attttctcct tacgcatctg tgcggtattt cacaccgcat acgtcaaagc aaccatagta
     3841 cgcgccctgt agcggcgcat taagcgcggc gggtgtggtg gttacgcgca gcgtgaccgc
     3901 tacacttgcc agcgccttag cgcccgctcc tttcgctttc ttcccttcct ttctcgccac
     3961 gttcgccggc tttccccgtc aagctctaaa tcgggggctc cctttagggt tccgatttag
     4021 tgctttacgg cacctcgacc ccaaaaaact tgatttgggt gatggttcac gtagtgggcc
     4081 atcgccctga tagacggttt ttcgcccttt gacgttggag tccacgttct ttaatagtgg
     4141 actcttgttc caaactggaa caacactcaa ctctatctcg ggctattctt ttgatttata
     4201 agggattttg ccgatttcgg tctattggtt aaaaaatgag ctgatttaac aaaaatttaa
     4261 cgcgaatttt aacaaaatat taacgtttac aattttatgg tgcactctca gtacaatctg
     4321 ctctgatgcc gcatagttaa gccagccccg acacccgcca acacccgctg acgcgccctg
     4381 acgggcttgt ctgctcccgg catccgctta cagacaagct gtgaccgtct ccgggagctg
     4441 catgtgtcag aggttttcac cgtcatcacc gaaacgcgcg agacgaaagg gcctcgtgat
     4501 acgcctattt ttataggtta atgtcatgat aataatggtt tcttagacgt caggtggcac
     4561 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     4621 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     4681 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     4741 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     4801 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     4861 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     4921 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     4981 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     5041 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     5101 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     5161 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     5221 gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     5281 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     5341 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     5401 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     5461 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     5521 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     5581 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     5641 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     5701 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     5761 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     5821 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     5881 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     5941 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     6001 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     6061 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     6121 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     6181 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     6241 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     6301 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgctggcctt ttgctcacat
     6361 gtcctgcagg cagctgcgcg ctcgctcgct cactgaggcc gcccgggcgt cgggcgacct
     6421 ttggtcgccc ggcctcagtg agcgagcgag cgcgcagaga gggagtggcc aactccatca
     6481 ctaggggttc ctgcggccgc aaacttgttt attgcagctt ataatggtta caaataaagc
     6541 aatagcatca caaatttcac aaataaagca tttttttcac tgcattctag ttgtggtttg
     6601 tccaaactca tcaatgtatc ttatcatgtc tgagatcctt acttagttac ccggggagca
     6661 tgtcaaggtc aaaatcgtca agagcgtcag caggcagcat atcaaggtca aagtcgtcaa
     6721 gggcatcggc tgggagcatg tctaagtcaa aatcgtcaag ggcgtcggcc ggcccgccgc
     6781 tttcgcactt tagctgtttc tccaggccac atatgattag ttccaggccg aaaaggaagg
     6841 caggttcggc tccctgatgg tcgaacagct caattgcttg tctcagaagt gggggcatag
     6901 a
//
