LOCUS       sequence-379367-        7522 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV-vector for bidirectional optogenetic manipulation of 
            tTA-positive neurons.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7522)
  AUTHORS   Wiegert Lab
  TITLE     Wiegert Lab Plasmids 2022
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7522)
  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            "Unpublished"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: TRE3GV-somBiPOLES-mKate2
FEATURES             Location/Qualifiers
     source          1..7522
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      complement(182..770)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(262..281)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(941..1732,1733..1801))
                     /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     1564..1583
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(1802..1906)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(1932..2387)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(2069..2090)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     2281..2300
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   2459..2588
                     /label=AAV2 ITR
     promoter        2680..3029
                     /label=TRE3GV promoter
                     /note="3rd-generation Tet-responsive promoter that can be 
                     activated by binding of Tet-On(R) 3G, optimized for 
                     retroviral and lentiviral vectors"
     protein_bind    2688..2706
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2724..2742
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2760..2778
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2796..2814
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2832..2850
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2868..2886
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    2904..2922
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     2968..2992
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     CDS             4010..4705
                     /codon_start=1
                     /product="monomeric far-red fluorescent protein (Shcherbo 
                     et al., 2009)"
                     /label=mKate2
                     /note="mammalian codon-optimized"
                     /translation="MVSELIKENMHMKLYMEGTVNNHHFKCTSEGEGKPYEGTQTMRIK
                     AVEGGPLPFAFDILATSFMYGSKTFINHTQGIPDFFKQSFPEGFTWERVTTYEDGGVLT
                     ATQDTSLQDGCLIYNVKIRGVNFPSNGPVMQKKTLGWEASTETLYPADGGLEGRADMAL
                     KLVGGGHLICNLKTTYRSKKPAKNLKMPGVYYVDRRLERIKEADKETYVEQHEVAVARY
                     CDLPSKLGHR"
     CDS             5066..6109
                     /codon_start=1
                     /product="red light-drivable channelrhodopsin from 
                     Chlamydomonas noctigama (Klapoetke et al., 2014)"
                     /label=Chrimson
                     /translation="ELISSATRSLFAAGGINPWPNPYHHEDMGCGGMTPTGECFSTEWW
                     CDPSYGLSDAGYGYCFVEATGGYLVVGVEKKQAWLHSRGTPGEKIGAQVCQWIAFSIAI
                     ALLTFYGFSAWKATCGWEEVYVCCVEVLFVTLEIFKEFSSPATVYLSTGNHAYCLRYFE
                     WLLSCPVILIKLSNLSGLKNDYSKRTMGLIVSCVGMIVFGMAAGLATDWLKWLLYIVSC
                     IYGGYMYFQAAKCYVEANHSVPKGHCRMVVKLMAYAYFASWGSYPILWAVGPEGLLKLS
                     PYANSIGHSICDIIAKEFWTFLAHHLRIKIHEHILIHGDIRKTTKMEIGGEEVEVEEFV
                     EEEDEDTV"
     misc_feature    6332..6920
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(6385..6405)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(6803..6814)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    6961..7168
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     repeat_region   7338..7467
                     /label=AAV2 ITR
ORIGIN
        1 cttccgcttc ctcgctcact gactcgctgc gctcggtcgt tcggctgcgg cgagcggtat
       61 cagctcactc aaaggcggta atacggttat ccacagaatc aggggataac gcaggaaaga
      121 acatgtgagc aaaaggccag caaaaggcca ggaaccgtaa aaaggccgcg ttgctggcgt
      181 ttttccatag gctccgcccc cctgacgagc atcacaaaaa tcgacgctca agtcagaggt
      241 ggcgaaaccc gacaggacta taaagatacc aggcgtttcc ccctggaagc tccctcgtgc
      301 gctctcctgt tccgaccctg ccgcttaccg gatacctgtc cgcctttctc ccttcgggaa
      361 gcgtggcgct ttctcatagc tcacgctgta ggtatctcag ttcggtgtag gtcgttcgct
      421 ccaagctggg ctgtgtgcac gaaccccccg ttcagcccga ccgctgcgcc ttatccggta
      481 actatcgtct tgagtccaac ccggtaagac acgacttatc gccactggca gcagccactg
      541 gtaacaggat tagcagagcg aggtatgtag gcggtgctac agagttcttg aagtggtggc
      601 ctaactacgg ctacactaga agaacagtat ttggtatctg cgctctgctg aagccagtta
      661 ccttcggaaa aagagttggt agctcttgat ccggcaaaca aaccaccgct ggtagcggtg
      721 gtttttttgt ttgcaagcag cagattacgc gcagaaaaaa aggatctcaa gaagatcctt
      781 tgatcttttc tacggggtct gacgctcagt ggaacgaaaa ctcacgttaa gggattttgg
      841 tcatgagatt atcaaaaagg atcttcacct agatcctttt aaattaaaaa tgaagtttta
      901 aatcaatcta aagtatatat gagtaaactt ggtctgacag ttaccaatgc ttaatcagtg
      961 aggcacctat ctcagcgatc tgtctatttc gttcatccat agttgcctga ctccccgtcg
     1021 tgtagataac tacgatacgg gagggcttac catctggccc cagtgctgca atgataccgc
     1081 gagacccacg ctcaccggct ccagatttat cagcaataaa ccagccagcc ggaagggccg
     1141 agcgcagaag tggtcctgca actttatccg cctccatcca gtctattaat tgttgccggg
     1201 aagctagagt aagtagttcg ccagttaata gtttgcgcaa cgttgttgcc attgctacag
     1261 gcatcgtggt gtcacgctcg tcgtttggta tggcttcatt cagctccggt tcccaacgat
     1321 caaggcgagt tacatgatcc cccatgttgt gcaaaaaagc ggttagctcc ttcggtcctc
     1381 cgatcgttgt cagaagtaag ttggccgcag tgttatcact catggttatg gcagcactgc
     1441 ataattctct tactgtcatg ccatccgtaa gatgcttttc tgtgactggt gagtactcaa
     1501 ccaagtcatt ctgagaatag tgtatgcggc gaccgagttg ctcttgcccg gcgtcaatac
     1561 gggataatac cgcgccacat agcagaactt taaaagtgct catcattgga aaacgttctt
     1621 cggggcgaaa actctcaagg atcttaccgc tgttgagatc cagttcgatg taacccactc
     1681 gtgcacccaa ctgatcttca gcatctttta ctttcaccag cgtttctggg tgagcaaaaa
     1741 caggaaggca aaatgccgca aaaaagggaa taagggcgac acggaaatgt tgaatactca
     1801 tactcttcct ttttcaatat tattgaagca tttatcaggg ttattgtctc atgagcggat
     1861 acatatttga atgtatttag aaaaataaac aaataggggt tccgcgcaca tttccccgaa
     1921 aagtgccacc taaattgtaa gcgttaatat tttgttaaaa ttcgcgttaa atttttgtta
     1981 aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata aatcaaaaga
     2041 atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac tattaaagaa
     2101 cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc cactacgtga
     2161 accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa atcggaaccc
     2221 taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg cgagaaagga
     2281 agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg tcacgctgcg
     2341 cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtccc attcgccatt
     2401 caggctgcgc aactgttggg aagggcgatc ggtgcgggcc tcttcgctat tacgccagct
     2461 gcgcgctcgc tcgctcactg aggccgcccg ggcaaagccc gggcgtcggg cgacctttgg
     2521 tcgcccggcc tcagtgagcg agcgagcgcg cagagaggga gtggccaact ccatcactag
     2581 gggttccttg tagttaatga ttaacccgcc atgctactta tctacgtagc catgctctag
     2641 gaagatcgta ggttcgatga ggccctttcg tcttcactcg agtttactcc ctatcagtga
     2701 tagagaacgt atgaagagtt tactccctat cagtgataga gaacgtatgc agactttact
     2761 ccctatcagt gatagagaac gtataaggag tttactccct atcagtgata gagaacgtat
     2821 gaccagttta ctccctatca gtgatagaga acgtatctac agtttactcc ctatcagtga
     2881 tagagaacgt atatccagtt tactccctat cagtgataga gaacgtatgt cgaggtaggc
     2941 gtgtacggtg ggcgcctata aaagcagagc tcgtttagtg aaccgtcaga tcgcctggag
     3001 caattccaca acacttttgt cttatacttg gatccgggcc cgcggccgat ccatggcatc
     3061 acaggtcgtc tatggggaat gggcatcaac tcacaccgag tgctacaata tgtccagaat
     3121 cgactccacc ttcgtctcac tgctgcagct ggtgtgggcc gtggtctcag gctgccagac
     3181 aatcttcatg attagcaggg ctcccaaggt cccttgggag tccgtgtacc tgccctttgt
     3241 cgaatctatc acttatgcac tggccagtac tggcaacgga accctgcaga tgagggacgg
     3301 cagattcttt ccttggagcc gaatggcctc ctggctgtgc acctgtccaa tcatgctggg
     3361 acagatttct aacatggctc tggtgaagta caaaagtatc ccactgaatc ccattgcaca
     3421 ggccgcttcc atcattagag tggtcatggg aatcaccgct acaatttctc ccgcagagta
     3481 tatgaagtgg ctgttctttt tctttggggc cacctgcctg gtgttcgagt acagtgtggt
     3541 cttcacaatc tttcaagtgg ggctgtatgg ctttgaatct gtcgggacac ctctggcaca
     3601 gaaggtggtg gtgaggatca agatgctgcg cctgatcttc tttattgctt ggactatgtt
     3661 cccaatcgtg tggctgattt cacccaccgg agtctgcgtg atccacgaga acgtgagcgc
     3721 cattctgtac ctgctggctg acggcctgtg caaaaataca tatggagtga tcctgtggag
     3781 cactgcctgg ggagtgctgg aagggaagtg ggaccccgca tgtctgccag ggcaggagaa
     3841 acccgaagcc gacgatcctt ttggcctgaa tcacgagaag aatgcccctc ctaacgacga
     3901 agtcaatatc agaatgtttg gaaggggacg tacggcggcc aagagcagga tcaccagcga
     3961 gggcgagtac atccccctgg accagatcga catcaacgtg gcggtaccca tggtgagcga
     4021 gctgattaag gagaacatgc acatgaagct gtacatggag ggcaccgtga acaaccacca
     4081 cttcaagtgc acatccgagg gcgaaggcaa gccctacgag ggcacccaga ccatgagaat
     4141 caaggcggtc gagggcggcc ctctcccctt cgccttcgac atcctggcta ccagcttcat
     4201 gtacggcagc aaaaccttca tcaaccacac ccagggcatc cccgacttct ttaagcagtc
     4261 cttccccgag ggcttcacat gggagagagt caccacatac gaagacgggg gcgtgctgac
     4321 cgctacccag gacaccagcc tccaggacgg ctgcctcatc tacaacgtca agatcagagg
     4381 ggtgaacttc ccatccaacg gccctgtgat gcagaagaaa acactcggct gggaggcctc
     4441 caccgagacc ctgtaccccg ctgacggcgg cctggaaggc agagccgaca tggccctgaa
     4501 gctcgtgggc gggggccacc tgatctgcaa cttgaagacc acatacagat ccaagaaacc
     4561 cgctaagaac ctcaagatgc ccggcgtcta ctatgtggac agaagactgg aaagaatcaa
     4621 ggaggccgac aaagagacct acgtcgagca gcacgaggtg gctgtggcca gatactgcga
     4681 cctccctagc aaactggggc acagaaccgg tgcagccctg caggagaaga agtcatgcag
     4741 ccagcgcatg gccgaattcc ggcaatactg ttggaacccg gacactgggc agatgctggg
     4801 ccgcacccca gcccggtggg tgtggatcag cctgtactat gcagctttct acgtggtcat
     4861 gactgggctc tttgccttgt gcatctatgt gctgatgcag accattgatc cctacacccc
     4921 cgactaccag gaccagttaa agtcaccggg ggtaaccttg agaccggatg tgtatgggga
     4981 aagagggctg cagatttcct acaacatctc tgaaaacagc tctagacagg cccagatcac
     5041 cggacgtccg gagggagctg tacaagagct gatcagcagc gccaccagat ctctgtttgc
     5101 cgccggaggc atcaaccctt ggcctaaccc ctaccaccac gaggacatgg gctgtggagg
     5161 aatgacacct acaggcgagt gcttcagcac cgagtggtgg tgtgaccctt cttacggact
     5221 gagcgacgcc ggatacggat attgcttcgt ggaggccaca ggcggctacc tggtcgtggg
     5281 agtggagaag aagcaggctt ggctgcacag cagaggcaca ccaggagaaa agatcggcgc
     5341 ccaggtctgc cagtggattg ctttcagcat cgccatcgcc ctgctgacat tctacggctt
     5401 cagcgcttgg aaggccactt gcggttggga ggaggtctac gtctgttgcg tcgaggtgct
     5461 gttcgtgacc ctggagatct tcaaggagtt cagcagcccc gccacagtgt acctgtctac
     5521 cggcaaccac gcctattgcc tgcgctactt cgagtggctg ctgtcttgcc ccgtgatcct
     5581 gatcaagctg agcaacctga gcggcctgaa gaacgactac agcaagcgga ccatgggcct
     5641 gatcgtgtct tgcgtgggaa tgatcgtgtt cggcatggcc gcaggactgg ctaccgattg
     5701 gctcaagtgg ctgctgtata tcgtgtcttg catctacggc ggctacatgt acttccaggc
     5761 cgccaagtgc tacgtggaag ccaaccacag cgtgcctaaa ggccattgcc gcatggtcgt
     5821 gaagctgatg gcctacgctt acttcgcctc ttggggcagc tacccaatcc tctgggcagt
     5881 gggaccagaa ggactgctga agctgagccc ttacgccaac agcatcggcc acagcatctg
     5941 cgacatcatc gccaaggagt tttggacctt cctggcccac cacctgagga tcaagatcca
     6001 cgagcacatc ctgatccacg gcgacatccg gaagaccacc aagatggaga tcggaggcga
     6061 ggaggtggaa gtggaagagt tcgtggagga ggaggacgag gacacagtgc tcgagcagag
     6121 ccagcccatc ctgaacacca aggagatggc cccccagagc aagccccccg aggagctgga
     6181 gatgagcagc atgcccagcc ccgtggcccc cctgcccgcc cgcaccgagg gcgtgatcga
     6241 catgcgcagc atgagcagca tcgacagctt catcagctgc gccaccgact tccccgaggc
     6301 cacccgcttc tgatctagaa agcttatcga taatcaacct ctggattaca aaatttgtga
     6361 aagattgact ggtattctta actatgttgc tccttttacg ctatgtggat acgctgcttt
     6421 aatgcctttg tatcatgcta ttgcttcccg tatggctttc attttctcct ccttgtataa
     6481 atcctggttg ctgtctcttt atgaggagtt gtggcccgtt gtcaggcaac gtggcgtggt
     6541 gtgcactgtg tttgctgacg caacccccac tggttggggc attgccacca cctgtcagct
     6601 cctttccggg actttcgctt tccccctccc tattgccacg gcggaactca tcgccgcctg
     6661 ccttgcccgc tgctggacag gggctcggct gttgggcact gacaattccg tggtgttgtc
     6721 ggggaaatca tcgtcctttc cttggctgct cgcctgtgtt gccacctgga ttctgcgcgg
     6781 gacgtccttc tgctacgtcc cttcggccct caatccagcg gaccttcctt cccgcggcct
     6841 gctgccggct ctgcggcctc ttccgcgtct tcgccttcgc cctcagacga gtcggatctc
     6901 cctttgggcc gcctccccgc atcgataccg tcgacccggg cggccgctca taaagctcga
     6961 ctgtgccttc tagttgccag ccatctgttg tttgcccctc ccccgtgcct tccttgaccc
     7021 tggaaggtgc cactcccact gtcctttcct aataaaatga ggaaattgca tcgcattgtc
     7081 tgagtaggtg tcattctatt ctggggggtg gggtggggca ggacagcaag ggggaggatt
     7141 gggaagacaa tagcaggcat gctggggatg cggtccggac tgtacaattg cattcatttt
     7201 atgtttcagg ttcaggggga gatgtgggag gttttttaaa gcaagtaaaa cctctacaaa
     7261 tgtggtaaaa tcgataagga tcttcctaga gcatggctac gtagataagt agcatggcgg
     7321 gttaatcatt aactacaagg aacccctagt gatggagttg gccactccct ctctgcgcgc
     7381 tcgctcgctc actgaggccg ggcgaccaaa ggtcgcccga cgcccgggct ttgcccgggc
     7441 ggcctcagtg agcgagcgag cgcgcagctg cattaatgaa tcggccaacg cgcggggaga
     7501 ggcggtttgc gtattgggcg ct
//
