LOCUS       sequence-486971-        6901 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Bidirectional AAV transfer plasmid expressing hSyn-tTA and 
            TRE-FLEX-mNeonGreen-MYC 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  JOURNAL   Unpublished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            "Unpublished"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-MYC
FEATURES             Location/Qualifiers
     source          1..6901
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(107..126)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      920..1508
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     1409..1428
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   1570..1699
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     polyA_signal    1709..1830
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1746..1765)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1800..1819
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     CDS             complement(join(1854..1862,1863..1898,1899..1901,1902..1937,
                     1938..1940,1941..1976,1977..1982,1983..2600))
                     /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"
     protein_bind    3056..3074
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3093..3111
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3130..3148
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3167..3185
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3204..3222
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3241..3259
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3278..3296
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3315..3333
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     3355..3379
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    complement(3478..3511)
                     /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(3562..3595)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(3702..3731)
                     /codon_start=1
                     /product="Myc (human c-Myc proto-oncogene) epitope tag"
                     /label=Myc
                     /translation="EQKLISEEDL"
     regulatory      4178..4187
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      4439..4448
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    4455..4488
                     /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    4539..4572
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4597..5185
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4650..4670)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(5068..5079)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    5217..5693
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(5416..5435)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   5733..5873
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5733..5862
                     /label=AAV2 ITR
     rep_origin      5948..6403
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6035..6054)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6245..6266
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(6420..6439)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     6539..6561
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(6599..6617)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        6685..6789
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6790..6858,6859..6901,1..749)
                     /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"
ORIGIN
        1 tgggtgcacg agtgggttac atcgaactgg atctcaacag cggtaagatc cttgagagtt
       61 ttcgccccga agaacgtttt ccaatgatga gcacttttaa agttctgcta tgtggcgcgg
      121 tattatcccg tattgacgcc gggcaagagc aactcggtcg ccgcatacac tattctcaga
      181 atgacttggt tgagtactca ccagtcacag aaaagcatct tacggatggc atgacagtaa
      241 gagaattatg cagtgctgcc ataaccatga gtgataacac tgcggccaac ttacttctga
      301 caacgatcgg aggaccgaag gagctaaccg cttttttgca caacatgggg gatcatgtaa
      361 ctcgccttga tcgttgggaa ccggagctga atgaagccat accaaacgac gagcgtgaca
      421 ccacgatgcc tgtagcaatg gcaacaacgt tgcgcaaact attaactggc gaactactta
      481 ctctagcttc ccggcaacaa ttaatagact ggatggaggc ggataaagtt gcaggaccac
      541 ttctgcgctc ggcccttccg gctggctggt ttattgctga taaatctgga gccggtgagc
      601 gtgggtctcg cggtatcatt gcagcactgg ggccagatgg taagccctcc cgtatcgtag
      661 ttatctacac gacggggagt caggcaacta tggatgaacg aaatagacag atcgctgaga
      721 taggtgcctc actgattaag cattggtaac tgtcagacca agtttactca tatatacttt
      781 agattgattt aaaacttcat ttttaattta aaaggatcta ggtgaagatc ctttttgata
      841 atctcatgac caaaatccct taacgtgagt tttcgttcca ctgagcgtca gaccccgtag
      901 aaaagatcaa aggatcttct tgagatcctt tttttctgcg cgtaatctgc tgcttgcaaa
      961 caaaaaaacc accgctacca gcggtggttt gtttgccgga tcaagagcta ccaactcttt
     1021 ttccgaaggt aactggcttc agcagagcgc agataccaaa tactgttctt ctagtgtagc
     1081 cgtagttagg ccaccacttc aagaactctg tagcaccgcc tacatacctc gctctgctaa
     1141 tcctgttacc agtggctgct gccagtggcg ataagtcgtg tcttaccggg ttggactcaa
     1201 gacgatagtt accggataag gcgcagcggt cgggctgaac ggggggttcg tgcacacagc
     1261 ccagcttgga gcgaacgacc tacaccgaac tgagatacct acagcgtgag ctatgagaaa
     1321 gcgccacgct tcccgaaggg agaaaggcgg acaggtatcc ggtaagcggc agggtcggaa
     1381 caggagagcg cacgagggag cttccagggg gaaacgcctg gtatctttat agtcctgtcg
     1441 ggtttcgcca cctctgactt gagcgtcgat ttttgtgatg ctcgtcaggg gggcggagcc
     1501 tatggaaaaa cgccagcaac gcggcctttt tacggttcct ggccttttgc tggccttttg
     1561 ctcacatgtc ctgcaggcag ctgcgcgctc gctcgctcac tgaggccgcc cgggcgtcgg
     1621 gcgacctttg gtcgcccggc ctcagtgagc gagcgagcgc gcagagaggg agtggccaac
     1681 tccatcacta ggggttcctg cggccgcaaa cttgtttatt gcagcttata atggttacaa
     1741 ataaagcaat agcatcacaa atttcacaaa taaagcattt ttttcactgc attctagttg
     1801 tggtttgtcc aaactcatca atgtatctta tcatgtctga gatccttact tagttacccg
     1861 gggagcatgt caaggtcaaa atcgtcaaga gcgtcagcag gcagcatatc aaggtcaaag
     1921 tcgtcaaggg catcggctgg gagcatgtct aagtcaaaat cgtcaagggc gtcggccggc
     1981 ccgccgcttt cgcactttag ctgtttctcc aggccacata tgattagttc caggccgaaa
     2041 aggaaggcag gttcggctcc ctgatggtcg aacagctcaa ttgcttgtct cagaagtggg
     2101 ggcatagaat cggtggtagg tgtctctctt tcctcttttg ctacttgatg ctcctgatcc
     2161 tccaatacgc agcccagtgt aaagtggccc acggcggaca gagcgtacag tgcgttctcc
     2221 agggagaagc cttgctgaca caggaacgcg agctgatttt ccagggtttc gtactgtttc
     2281 tctgttgggc gggtgccgag atgcacttta gccccgtcgc gatgtgagag gagagcacag
     2341 cggaatgact tggcgttgtt ccgcagaaag tcttgccatg actcgccttc cagggggcag
     2401 aagtgggtat gatgcctgtc cagcatctcg attgccaggg catcgagcag ggcccgcttg
     2461 ttcttcacgt gccagtacag ggtaggctgc tcaactccca gcttttgagc gagtttcctt
     2521 gtcgtcaggc cttcgatacc gacttcattg agtaattcca gagcagagtt tatgactttg
     2581 ctcttgtcca gtctagacat ggtgaattcg atatcaagct tatcgatacc gtccgccgca
     2641 gcgcagatgg tcgcgcccgt gcccccctat ctcgcgcctc gcgtggtgcg gtccggctgg
     2701 gccggcggcg gcgcggacgc gaccaaggtg gccgggaagg ggagtttgcg ggggaccggc
     2761 gagtgacgtc agcgcgcctt cagtgctgag gcggcggtgg cgcgcgccgc caggcggggg
     2821 cgaaggcact gtccgcggtg ctgaagctgg cagtgcgcac gcgcctcgcc gcatcctgtt
     2881 tcccctcccc ctctctgata ggggatgcgc aatttgggga atgggggttg ggtgcttgtc
     2941 cagtgggtcg gggtcggtcg tcaggtaggc acccccaccc cgcctcatcc tggtcctaaa
     3001 acccacttgc actcatacgc agggccctct gcagtatcaa gtgccacctg acgtctccct
     3061 atcagtgata gagaagtcga cacgtctcga gctccctatc agtgatagag aaggtacgtc
     3121 tagaacgtct ccctatcagt gatagagaag tcgacacgtc tcgagctccc tatcagtgat
     3181 agagaaggta cgtctagaac gtctccctat cagtgataga gaagtcgaca cgtctcgagc
     3241 tccctatcag tgatagagaa ggtacgtcta gaacgtctcc ctatcagtga tagagaagtc
     3301 gacacgtctc gagctcccta tcagtgatag agaagctacc ccctatataa gcagagctcg
     3361 tttagtgaac cgtcagatcg cctggagacg ccatccacgc tgttttgacc tccatagaag
     3421 acaccgggac cgatccagcc tggatccggt accggatcct ctagagtcga ctccggaata
     3481 acttcgtata ggatacttta tacgaagtta tgcagaatgg tagctggatt gtagctgcta
     3541 ttagcaatat gaaacctctt aataacttcg tatagcatac attatacgaa gttatggcgc
     3601 gcctcacttg tccaaggaat tcttactttg attatcgata agcttgacct ggataacgct
     3661 gggcgcaagt tgatcagacc agtcctactg aagaattctt acaggtcctc ctcgctgatc
     3721 agcttctgct cgctgccgcc cagctcgtcc atgcccatca catcggtaaa ggccttttgc
     3781 cactccttga agttgagctc ggtcttggag tgcttgagct ccgtcttacg gaacacgtac
     3841 atcggctggt tcttcagata gttagccgcc attggcttgg caaaggtgta ggtggtccgc
     3901 gcagtgctcc ggtagcgctt gccatttcca gtggtgtaac tccacttaaa ggtactgatg
     3961 atggttttgt cgttggggta agtcttcttc gacctgcacc agtccgcagc ggtcagcgag
     4021 ttggtcatca caggaccgtc agcagggaaa ccagtcccct tcacctgggc ctctcctttg
     4081 atgtggcttc cctcgtaggt gtagcggtag ttaacagtaa gggaggcacc atcttcaaac
     4141 tgcattgtgc gatggacttg gtatccggag ccatctacca tggcggcctg gaaaggcgac
     4201 atcccgtcag ggtagggcag gtactgatgg aagccatacc cgatatgagg gaccagaatc
     4261 cagggggaga actggaggtc acccttggtg gacttcaggt ttaactcctc ataaccatca
     4321 tttggattgc cggtgccctg acccaccatg tcaaagtcca caccgttgat ggagccaaag
     4381 atgtgtaact catgtgtcgc tgggagagag gccatgttat cctcctcgcc cttgctcacc
     4441 atggtggcgc tagcataact tcgtataaag tatcctatac gaagttattt gccttaaccc
     4501 agaaattatc actgttattc tttagaatgg tgcaaagaat aacttcgtat aatgtatgct
     4561 atacgaagtt atgaattcga tatcaagctt atcgataatc aacctctgga ttacaaaatt
     4621 tgtgaaagat tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct
     4681 gctttaatgc ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg
     4741 tataaatcct ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc
     4801 gtggtgtgca ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt
     4861 cagctccttt ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc
     4921 gcctgccttg cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg
     4981 ttgtcgggga aatcatcgtc ctttccttgg ctgctcgcct atgttgccac ctggattctg
     5041 cgcgggacgt ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc
     5101 ggcctgctgc cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg
     5161 atctcccttt gggccgcctc cccgcatcga taccgagcgc tgctcgagag atctacgggt
     5221 ggcatccctg tgacccctcc ccagtgcctc tcctggccct ggaagttgcc actccagtgc
     5281 ccaccagcct tgtcctaata aaattaagtt gcatcatttt gtctgactag gtgtccttct
     5341 ataatattat ggggtggagg ggggtggtat ggagcaaggg gcaagttggg aagacaacct
     5401 gtagggcctg cggggtctat tgggaaccaa gctggagtgc agtggcacaa tcttggctca
     5461 ctgcaatctc cgcctcctgg gttcaagcga ttctcctgcc tcagcctccc gagttgttgg
     5521 gattccaggc atgcatgacc aggctcagct aatttttgtt tttttggtag agacggggtt
     5581 tcaccatatt ggccaggctg gtctccaact cctaatctca ggtgatctac ccaccttggc
     5641 ctcccaaatt gctgggatta caggcgtgaa ccactgctcc cttccctgtc cttctgattt
     5701 tgtaggtaac cacgtgcgga ccgagcggcc gcaggaaccc ctagtgatgg agttggccac
     5761 tccctctctg cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc
     5821 gggctttgcc cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct
     5881 gatgcggtat tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa
     5941 ccatagtacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc
     6001 gtgaccgcta cacttgccag cgccttagcg cccgctcctt tcgctttctt cccttccttt
     6061 ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc
     6121 cgatttagtg ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt
     6181 agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt
     6241 aatagtggac tcttgttcca aactggaaca acactcaact ctatctcggg ctattctttt
     6301 gatttataag ggattttgcc gatttcggtc tattggttaa aaaatgagct gatttaacaa
     6361 aaatttaacg cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt
     6421 acaatctgct ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac
     6481 gcgccctgac gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc
     6541 gggagctgca tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc
     6601 ctcgtgatac gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca
     6661 ggtggcactt ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat
     6721 tcaaatatgt atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa
     6781 aggaagagta tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt
     6841 tgccttcctg tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag
     6901 t
//
