LOCUS       sequence-486973-        6904 bp    DNA     circular SYN 23-SEP-2025
DEFINITION  Bidirectional AAV transfer plasmid expressing hSyn-tTA and 
            TRE-FLEX-mNeonGreen-HSV in the other direction..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6904)
  AUTHORS   n/a
  TITLE     E11 Bio PRISM Collection Plasmids 2025
  JOURNAL   unpublished
REFERENCE   2  (bases 1 to 6904)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported Sep 23, 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            "unpublished"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-HSV
FEATURES             Location/Qualifiers
     source          1..6904
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     11..30
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(249..353)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     421..439
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     primer_bind     complement(477..499)
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     599..618
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     rep_origin      complement(635..1090)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(772..793)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     984..1003
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   1165..1305
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   1176..1305
                     /label=AAV2 ITR
     polyA_signal    1345..1821
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     1603..1622
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     misc_feature    1853..2441
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1959..1970
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     2368..2388
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    complement(2466..2499)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(2550..2583)
                     /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)."
     regulatory      2590..2599
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      2851..2860
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             3307..3339
                     /codon_start=1
                     /product="HSV (herpes simplex virus) epitope tag"
                     /label=HSV tag
                     /translation="QPELAPEDPED"
     protein_bind    3446..3479
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    3530..3563
                     /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)."
     primer_bind     complement(3662..3686)
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    3708..3726
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3745..3763
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3782..3800
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3819..3837
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3856..3874
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3893..3911
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3930..3948
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    3967..3985
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     CDS             join(4441..5058,5059..5064,5065..5100,5101..5103,5104..5139,
                     5140..5142,5143..5178,5179..5187)
                     /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"
     polyA_signal    5211..5332
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5222..5241)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5276..5295
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   5342..5471
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     rep_origin      complement(5533..6121)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5613..5632)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6292..6904,1..179,180..248))
                     /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 tcaatacggg ataataccgc gccacatagc agaactttaa aagtgctcat cattggaaaa
       61 cgttcttcgg ggcgaaaact ctcaaggatc ttaccgctgt tgagatccag ttcgatgtaa
      121 cccactcgtg cacccaactg atcttcagca tcttttactt tcaccagcgt ttctgggtga
      181 gcaaaaacag gaaggcaaaa tgccgcaaaa aagggaataa gggcgacacg gaaatgttga
      241 atactcatac tcttcctttt tcaatattat tgaagcattt atcagggtta ttgtctcatg
      301 agcggataca tatttgaatg tatttagaaa aataaacaaa taggggttcc gcgcacattt
      361 ccccgaaaag tgccacctga cgtctaagaa accattatta tcatgacatt aacctataaa
      421 aataggcgta tcacgaggcc ctttcgtctc gcgcgtttcg gtgatgacgg tgaaaacctc
      481 tgacacatgc agctcccgga gacggtcaca gcttgtctgt aagcggatgc cgggagcaga
      541 caagcccgtc agggcgcgtc agcgggtgtt ggcgggtgtc ggggctggct taactatgcg
      601 gcatcagagc agattgtact gagagtgcac cataaaattg taaacgttaa tattttgtta
      661 aaattcgcgt taaatttttg ttaaatcagc tcatttttta accaatagac cgaaatcggc
      721 aaaatccctt ataaatcaaa agaatagccc gagatagagt tgagtgttgt tccagtttgg
      781 aacaagagtc cactattaaa gaacgtggac tccaacgtca aagggcgaaa aaccgtctat
      841 cagggcgatg gcccactacg tgaaccatca cccaaatcaa gttttttggg gtcgaggtgc
      901 cgtaaagcac taaatcggaa ccctaaaggg agcccccgat ttagagcttg acggggaaag
      961 ccggcgaacg tggcgagaaa ggaagggaag aaagcgaaag gagcgggcgc taaggcgctg
     1021 gcaagtgtag cggtcacgct gcgcgtaacc accacacccg ccgcgcttaa tgcgccgcta
     1081 cagggcgcgt actatggttg ctttgacgta tgcggtgtga aataccgcac agatgcgtaa
     1141 ggagaaaata ccgcatcagg cgcccctgca ggcagctgcg cgctcgctcg ctcactgagg
     1201 ccgcccgggc aaagcccggg cgtcgggcga cctttggtcg cccggcctca gtgagcgagc
     1261 gagcgcgcag agagggagtg gccaactcca tcactagggg ttcctgcggc cgctcggtcc
     1321 gcacgtggtt acctacaaaa tcagaaggac agggaaggga gcagtggttc acgcctgtaa
     1381 tcccagcaat ttgggaggcc aaggtgggta gatcacctga gattaggagt tggagaccag
     1441 cctggccaat atggtgaaac cccgtctcta ccaaaaaaac aaaaattagc tgagcctggt
     1501 catgcatgcc tggaatccca acaactcggg aggctgaggc aggagaatcg cttgaaccca
     1561 ggaggcggag attgcagtga gccaagattg tgccactgca ctccagcttg gttcccaata
     1621 gaccccgcag gccctacagg ttgtcttccc aacttgcccc ttgctccata ccacccccct
     1681 ccaccccata atattataga aggacaccta gtcagacaaa atgatgcaac ttaattttat
     1741 taggacaagg ctggtgggca ctggagtggc aacttccagg gccaggagag gcactgggga
     1801 ggggtcacag ggatgccacc cgtagatctc tcgagcagcg ctcggtatcg atgcggggag
     1861 gcggcccaaa gggagatccg actcgtctga gggcgaaggc gaagacgcgg aagaggccgc
     1921 agagccggca gcaggccgcg ggaaggaagg tccgctggat tgagggccga agggacgtag
     1981 cagaaggacg tcccgcgcag aatccaggtg gcaacatagg cgagcagcca aggaaaggac
     2041 gatgatttcc ccgacaacac cacggaattg tcagtgccca acagccgagc ccctgtccag
     2101 cagcgggcaa ggcaggcggc gatgagttcc gccgtggcaa tagggagggg gaaagcgaaa
     2161 gtcccggaaa ggagctgaca ggtggtggca atgccccaac cagtgggggt tgcgtcagca
     2221 aacacagtgc acaccacgcc acgttgcctg acaacgggcc acaactcctc ataaagagac
     2281 agcaaccagg atttatacaa ggaggagaaa atgaaagcca tacgggaagc aatagcatga
     2341 tacaaaggca ttaaagcagc gtatccacat agcgtaaaag gagcaacata gttaagaata
     2401 ccagtcaatc tttcacaaat tttgtaatcc agaggttgat tatcgataag cttgatatcg
     2461 aattcataac ttcgtatagc atacattata cgaagttatt ctttgcacca ttctaaagaa
     2521 taacagtgat aatttctggg ttaaggcaaa taacttcgta taggatactt tatacgaagt
     2581 tatgctagcg ccaccatggt gagcaagggc gaggaggata acatggcctc tctcccagcg
     2641 acacatgagt tacacatctt tggctccatc aacggtgtgg actttgacat ggtgggtcag
     2701 ggcaccggca atccaaatga tggttatgag gagttaaacc tgaagtccac caagggtgac
     2761 ctccagttct ccccctggat tctggtccct catatcgggt atggcttcca tcagtacctg
     2821 ccctaccctg acgggatgtc gcctttccag gccgccatgg tagatggctc cggataccaa
     2881 gtccatcgca caatgcagtt tgaagatggt gcctccctta ctgttaacta ccgctacacc
     2941 tacgagggaa gccacatcaa aggagaggcc caggtgaagg ggactggttt ccctgctgac
     3001 ggtcctgtga tgaccaactc gctgaccgct gcggactggt gcaggtcgaa gaagacttac
     3061 cccaacgaca aaaccatcat cagtaccttt aagtggagtt acaccactgg aaatggcaag
     3121 cgctaccgga gcactgcgcg gaccacctac acctttgcca agccaatggc ggctaactat
     3181 ctgaagaacc agccgatgta cgtgttccgt aagacggagc tcaagcactc caagaccgag
     3241 ctcaacttca aggagtggca aaaggccttt accgatgtga tgggcatgga cgagctgggc
     3301 ggtagccagc cagaattggc ccccgaagac cctgaggact aagaattctt cagtaggact
     3361 ggtcgctgtc caaaggttct tcgtaacgcc aggtcaagct tatcgataat caaagtaaga
     3421 attccttgga caagtgaggc gcgccataac ttcgtataat gtatgctata cgaagttatt
     3481 aagaggtttc atattgctaa tagcagctac aatccagcta ccattctgca taacttcgta
     3541 taaagtatcc tatacgaagt tattccggag tcgactctag aggatccggt accggatcca
     3601 ggctggatcg gtcccggtgt cttctatgga ggtcaaaaca gcgtggatgg cgtctccagg
     3661 cgatctgacg gttcactaaa cgagctctgc ttatataggg ggtagcttct ctatcactga
     3721 tagggagctc gagacgtgtc gacttctcta tcactgatag ggagacgttc tagacgtacc
     3781 ttctctatca ctgataggga gctcgagacg tgtcgacttc tctatcactg atagggagac
     3841 gttctagacg taccttctct atcactgata gggagctcga gacgtgtcga cttctctatc
     3901 actgataggg agacgttcta gacgtacctt ctctatcact gatagggagc tcgagacgtg
     3961 tcgacttctc tatcactgat agggagacgt caggtggcac ttgatactgc agagggccct
     4021 gcgtatgagt gcaagtgggt tttaggacca ggatgaggcg gggtgggggt gcctacctga
     4081 cgaccgaccc cgacccactg gacaagcacc caacccccat tccccaaatt gcgcatcccc
     4141 tatcagagag ggggagggga aacaggatgc ggcgaggcgc gtgcgcactg ccagcttcag
     4201 caccgcggac agtgccttcg cccccgcctg gcggcgcgcg ccaccgccgc ctcagcactg
     4261 aaggcgcgct gacgtcactc gccggtcccc cgcaaactcc ccttcccggc caccttggtc
     4321 gcgtccgcgc cgccgccggc ccagccggac cgcaccacgc gaggcgcgag ataggggggc
     4381 acgggcgcga ccatctgcgc tgcggcggac ggtatcgata agcttgatat cgaattcacc
     4441 atgtctagac tggacaagag caaagtcata aactctgctc tggaattact caatgaagtc
     4501 ggtatcgaag gcctgacgac aaggaaactc gctcaaaagc tgggagttga gcagcctacc
     4561 ctgtactggc acgtgaagaa caagcgggcc ctgctcgatg ccctggcaat cgagatgctg
     4621 gacaggcatc atacccactt ctgccccctg gaaggcgagt catggcaaga ctttctgcgg
     4681 aacaacgcca agtcattccg ctgtgctctc ctctcacatc gcgacggggc taaagtgcat
     4741 ctcggcaccc gcccaacaga gaaacagtac gaaaccctgg aaaatcagct cgcgttcctg
     4801 tgtcagcaag gcttctccct ggagaacgca ctgtacgctc tgtccgccgt gggccacttt
     4861 acactgggct gcgtattgga ggatcaggag catcaagtag caaaagagga aagagagaca
     4921 cctaccaccg attctatgcc cccacttctg agacaagcaa ttgagctgtt cgaccatcag
     4981 ggagccgaac ctgccttcct tttcggcctg gaactaatca tatgtggcct ggagaaacag
     5041 ctaaagtgcg aaagcggcgg gccggccgac gcccttgacg attttgactt agacatgctc
     5101 ccagccgatg cccttgacga ctttgacctt gatatgctgc ctgctgacgc tcttgacgat
     5161 tttgaccttg acatgctccc cgggtaacta agtaaggatc tcagacatga taagatacat
     5221 tgatgagttt ggacaaacca caactagaat gcagtgaaaa aaatgcttta tttgtgaaat
     5281 ttgtgatgct attgctttat ttgtaaccat tataagctgc aataaacaag tttgcggccg
     5341 caggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag
     5401 gccgggcgac caaaggtcgc ccgacgcccg ggcggcctca gtgagcgagc gagcgcgcag
     5461 ctgcctgcag gacatgtgag caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc
     5521 gttgctggcg tttttccata ggctccgccc ccctgacgag catcacaaaa atcgacgctc
     5581 aagtcagagg tggcgaaacc cgacaggact ataaagatac caggcgtttc cccctggaag
     5641 ctccctcgtg cgctctcctg ttccgaccct gccgcttacc ggatacctgt ccgcctttct
     5701 cccttcggga agcgtggcgc tttctcatag ctcacgctgt aggtatctca gttcggtgta
     5761 ggtcgttcgc tccaagctgg gctgtgtgca cgaacccccc gttcagcccg accgctgcgc
     5821 cttatccggt aactatcgtc ttgagtccaa cccggtaaga cacgacttat cgccactggc
     5881 agcagccact ggtaacagga ttagcagagc gaggtatgta ggcggtgcta cagagttctt
     5941 gaagtggtgg cctaactacg gctacactag aagaacagta tttggtatct gcgctctgct
     6001 gaagccagtt accttcggaa aaagagttgg tagctcttga tccggcaaac aaaccaccgc
     6061 tggtagcggt ggtttttttg tttgcaagca gcagattacg cgcagaaaaa aaggatctca
     6121 agaagatcct ttgatctttt ctacggggtc tgacgctcag tggaacgaaa actcacgtta
     6181 agggattttg gtcatgagat tatcaaaaag gatcttcacc tagatccttt taaattaaaa
     6241 atgaagtttt aaatcaatct aaagtatata tgagtaaact tggtctgaca gttaccaatg
     6301 cttaatcagt gaggcaccta tctcagcgat ctgtctattt cgttcatcca tagttgcctg
     6361 actccccgtc gtgtagataa ctacgatacg ggagggctta ccatctggcc ccagtgctgc
     6421 aatgataccg cgagacccac gctcaccggc tccagattta tcagcaataa accagccagc
     6481 cggaagggcc gagcgcagaa gtggtcctgc aactttatcc gcctccatcc agtctattaa
     6541 ttgttgccgg gaagctagag taagtagttc gccagttaat agtttgcgca acgttgttgc
     6601 cattgctaca ggcatcgtgg tgtcacgctc gtcgtttggt atggcttcat tcagctccgg
     6661 ttcccaacga tcaaggcgag ttacatgatc ccccatgttg tgcaaaaaag cggttagctc
     6721 cttcggtcct ccgatcgttg tcagaagtaa gttggccgca gtgttatcac tcatggttat
     6781 ggcagcactg cataattctc ttactgtcat gccatccgta agatgctttt ctgtgactgg
     6841 tgagtactca accaagtcat tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc
     6901 ggcg
//
