LOCUS       sequence-329444-        5781 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes Flp-dependent rtTA.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5781)
  AUTHORS   Sabatini PV, Wang J, Rupp AC, Affinati AH, Flak JN, Li C, Olson DP, 
            Myers MG Jr
  TITLE     tTARGIT AAVs mediate the sensitive and flexible manipulation of 
            intersectional neuronal populations in mice.
  JOURNAL   eLife. 2021 Mar 11;10. pii: 66835. doi: 10.7554/eLife.66835.
  PUBMED    33704065
REFERENCE   2  (bases 1 to 5781)
  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: "eLife."; 
            date: "2021-03-11"; volume: "10. pii: 66835. doi"; pages: " 
            10.7554/eLife.66835"
COMMENT     Sequence Label: pAAV-hSYN1-fDIO-rtTA
FEATURES             Location/Qualifiers
     source          1..5781
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     promoter        160..607
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     protein_bind    647..665
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    683..701
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    804..837
                     /label=FRT3
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision with other FRT3 sites
                     (Bosch et al., 2015)"
     protein_bind    888..935
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             complement(join(970..978,979..1014,1015..1017,1018..1053,
                     1054..1056,1057..1092,1093..1098,1099..1716))
                     /codon_start=1
                     /product="improved tetracycline-controlled transactivator"
                     /label=rtTA-Advanced
                     /note="In the Tet-On(R) system, rtTA-Advanced binds to the 
                     Tet-responsive element and stimulates transcription only in
                     the presence of tetracycline or doxycycline."
                     /translation="MSRLDKSKVINGALELLNGVGIEGLTTRKLAQKLGVEQPTLYWHV
                     KNKRALLDALPIEMLDRHHTHFCPLEGESWQDFLRNNAKSFRCALLSHRDGAKVHLGTR
                     PTEKQYETLENQLAFLCQQGFSLENALYALSAVGHFTLGCVLEEQEHQVAKEERETPTT
                     DSMPPLLRQAIELFDRQGAEPAFLFGLELIICGLEKQLKCESGGPADALDDFDLDMLPA
                     DALDDFDLDMLPADALDDFDLDMLPG"
     protein_bind    complement(1727..1760)
                     /label=FRT3
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="supports FLP-mediated excision with other FRT3 sites
                     (Bosch et al., 2015)"
     protein_bind    complement(1825..1872)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     misc_feature    1893..2481
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1946..1966)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2364..2375)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2513..2989
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2712..2731)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   3029..3169
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   3029..3158
                     /label=AAV2 ITR
     rep_origin      3244..3699
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3331..3350)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3541..3562
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3716..3735)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3835..3857
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3895..3913)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3981..4085
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4086..4154,4155..4946)
                     /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(4304..4323)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5117..5705
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5606..5625
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   join(5767..5781,1..115)
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
ORIGIN
        1 gcgctcgctc gctcactgag gccgcccggg cgtcgggcga cctttggtcg cccggcctca
       61 gtgagcgagc gagcgcgcag agagggagtg gccaactcca tcactagggg ttcctgcggc
      121 cgcacgcgtg tgtctagact gcagagggcc ctgcgtatga gtgcaagtgg gttttaggac
      181 caggatgagg cggggtgggg gtgcctacct gacgaccgac cccgacccac tggacaagca
      241 cccaaccccc attccccaaa ttgcgcatcc cctatcagag agggggaggg gaaacaggat
      301 gcggcgaggc gcgtgcgcac tgccagcttc agcaccgcgg acagtgcctt cgcccccgcc
      361 tggcggcgcg cgccaccgcc gcctcagcac tgaaggcgcg ctgacgtcac tcgccggtcc
      421 cccgcaaact ccccttcccg gccaccttgg tcgcgtccgc gccgccgccg gcccagccgg
      481 accgcaccac gcgaggcgcg agataggggg gcacgggcgc gaccatctgc gctgcggcgc
      541 cggcgactca gcgctgcctc agtctgcggt gggcagcgga ggagtcgtgt cgtgcctgag
      601 agcgcagtcg agaacagatc tctaaaacag gtaagtccca ttaatctccc tatcagtgat
      661 agagaaggtc tgaagagttt actccctatc agtgatagag attaattcct ctactaacct
      721 tgttcatctt ttcttttttt ttctacaggt cctgggtgat taacagctta agggcccgga
      781 tccgatatcg aagttcctat tccgaagttc ctattcttca aatagtatag gaacttcgca
      841 gaatggtagc tggattgtag ctgctattag caatatgaaa cctcttagaa gttcctattc
      901 cgaagttcct attctctaga aagtatagga acttcgagct cggcgcgccg gaattcacta
      961 gtctcgagct tacccgggga gcatgtcaag gtcaaaatcg tcaagagcgt cagcaggcag
     1021 catatcaagg tcaaagtcgt caagggcatc ggctgggagc atgtctaagt caaaatcgtc
     1081 aagggcgtcg gccggcccgc cgctttcgca ctttagctgt ttctccaggc cacatatgat
     1141 tagttccagg ccgaaaagga aggcaggttc ggctccctgc cggtcgaaca gctcaattgc
     1201 ttgtctcaga agtgggggca tagaatcggt ggtaggtgtc tctctttcct cttttgctac
     1261 ttgatgctcc tgttcctcca atacgcagcc cagtgtaaag tggcccacgg cggacagagc
     1321 gtacagtgcg ttctccaggg agaagccttg ctgacacagg aacgcgagct gattttccag
     1381 ggtttcgtac tgtttctctg ttgggcgggt gccgagatgc actttagccc cgtcgcgatg
     1441 tgagaggaga gcacagcgga atgacttggc gttgttccgc agaaagtctt gccatgactc
     1501 gccttccagg gggcagaagt gggtatgatg cctgtccagc atctcgattg gcagggcatc
     1561 gagcagggcc cgcttgttct tcacgtgcca gtacagggta ggctgctcaa ctcccagctt
     1621 ttgagcgagt ttccttgtcg tcaggccttc gataccgact ccattgagta attccagagc
     1681 gccgtttatg actttgctct tgtccagtct agacatgtcg acccgggaag ttcctatact
     1741 atttgaagaa taggaacttc ggaataggaa cttcttgcct taacccagaa attatcactg
     1801 ttattcttta gaatggtgca aagagaagtt cctatacttt ctagagaata ggaacttcgg
     1861 aataggaact tcgaagtact aagcttatcg ataatcaacc tctggattac aaaatttgtg
     1921 aaagattgac tggtattctt aactatgttg ctccttttac gctatgtgga tacgctgctt
     1981 taatgccttt gtatcatgct attgcttccc gtatggcttt cattttctcc tccttgtata
     2041 aatcctggtt gctgtctctt tatgaggagt tgtggcccgt tgtcaggcaa cgtggcgtgg
     2101 tgtgcactgt gtttgctgac gcaaccccca ctggttgggg cattgccacc acctgtcagc
     2161 tcctttccgg gactttcgct ttccccctcc ctattgccac ggcggaactc atcgccgcct
     2221 gccttgcccg ctgctggaca ggggctcggc tgttgggcac tgacaattcc gtggtgttgt
     2281 cggggaaatc atcgtccttt ccttggctgc tcgcctatgt tgccacctgg attctgcgcg
     2341 ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc ggaccttcct tcccgcggcc
     2401 tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg ccctcagacg agtcggatct
     2461 ccctttgggc cgcctccccg catcgatacc gagcgctgct cgagagatct acgggtggca
     2521 tccctgtgac ccctccccag tgcctctcct ggccctggaa gttgccactc cagtgcccac
     2581 cagccttgtc ctaataaaat taagttgcat cattttgtct gactaggtgt ccttctataa
     2641 tattatgggg tggagggggg tggtatggag caaggggcaa gttgggaaga caacctgtag
     2701 ggcctgcggg gtctattggg aaccaagctg gagtgcagtg gcacaatctt ggctcactgc
     2761 aatctccgcc tcctgggttc aagcgattct cctgcctcag cctcccgagt tgttgggatt
     2821 ccaggcatgc atgaccaggc tcagctaatt tttgtttttt tggtagagac ggggtttcac
     2881 catattggcc aggctggtct ccaactccta atctcaggtg atctacccac cttggcctcc
     2941 caaattgctg ggattacagg cgtgaaccac tgctcccttc cctgtccttc tgattttgta
     3001 ggtaaccacg tgcggaccga gcggccgcag gaacccctag tgatggagtt ggccactccc
     3061 tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa aggtcgcccg acgcccgggc
     3121 tttgcccggg cggcctcagt gagcgagcga gcgcgcagct gcctgcaggg gcgcctgatg
     3181 cggtattttc tccttacgca tctgtgcggt atttcacacc gcatacgtca aagcaaccat
     3241 agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt ggtggttacg cgcagcgtga
     3301 ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc tttcttccct tcctttctcg
     3361 ccacgttcgc cggctttccc cgtcaagctc taaatcgggg gctcccttta gggttccgat
     3421 ttagtgcttt acggcacctc gaccccaaaa aacttgattt gggtgatggt tcacgtagtg
     3481 ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt ggagtccacg ttctttaata
     3541 gtggactctt gttccaaact ggaacaacac tcaactctat ctcgggctat tcttttgatt
     3601 tataagggat tttgccgatt tcggtctatt ggttaaaaaa tgagctgatt taacaaaaat
     3661 ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt atggtgcact ctcagtacaa
     3721 tctgctctga tgccgcatag ttaagccagc cccgacaccc gccaacaccc gctgacgcgc
     3781 cctgacgggc ttgtctgctc ccggcatccg cttacagaca agctgtgacc gtctccggga
     3841 gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg cgcgagacga aagggcctcg
     3901 tgatacgcct atttttatag gttaatgtca tgataataat ggtttcttag acgtcaggtg
     3961 gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt atttttctaa atacattcaa
     4021 atatgtatcc gctcatgaga caataaccct gataaatgct tcaataatat tgaaaaagga
     4081 agagtatgag tattcaacat ttccgtgtcg cccttattcc cttttttgcg gcattttgcc
     4141 ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa agatgctgaa gatcagttgg
     4201 gtgcacgagt gggttacatc gaactggatc tcaacagcgg taagatcctt gagagttttc
     4261 gccccgaaga acgttttcca atgatgagca cttttaaagt tctgctatgt ggcgcggtat
     4321 tatcccgtat tgacgccggg caagagcaac tcggtcgccg catacactat tctcagaatg
     4381 acttggttga gtactcacca gtcacagaaa agcatcttac ggatggcatg acagtaagag
     4441 aattatgcag tgctgccata accatgagtg ataacactgc ggccaactta cttctgacaa
     4501 cgatcggagg accgaaggag ctaaccgctt ttttgcacaa catgggggat catgtaactc
     4561 gccttgatcg ttgggaaccg gagctgaatg aagccatacc aaacgacgag cgtgacacca
     4621 cgatgcctgt agcaatggca acaacgttgc gcaaactatt aactggcgaa ctacttactc
     4681 tagcttcccg gcaacaatta atagactgga tggaggcgga taaagttgca ggaccacttc
     4741 tgcgctcggc ccttccggct ggctggttta ttgctgataa atctggagcc ggtgagcgtg
     4801 ggtctcgcgg tatcattgca gcactggggc cagatggtaa gccctcccgt atcgtagtta
     4861 tctacacgac ggggagtcag gcaactatgg atgaacgaaa tagacagatc gctgagatag
     4921 gtgcctcact gattaagcat tggtaactgt cagaccaagt ttactcatat atactttaga
     4981 ttgatttaaa acttcatttt taatttaaaa ggatctaggt gaagatcctt tttgataatc
     5041 tcatgaccaa aatcccttaa cgtgagtttt cgttccactg agcgtcagac cccgtagaaa
     5101 agatcaaagg atcttcttga gatccttttt ttctgcgcgt aatctgctgc ttgcaaacaa
     5161 aaaaaccacc gctaccagcg gtggtttgtt tgccggatca agagctacca actctttttc
     5221 cgaaggtaac tggcttcagc agagcgcaga taccaaatac tgttcttcta gtgtagccgt
     5281 agttaggcca ccacttcaag aactctgtag caccgcctac atacctcgct ctgctaatcc
     5341 tgttaccagt ggctgctgcc agtggcgata agtcgtgtct taccgggttg gactcaagac
     5401 gatagttacc ggataaggcg cagcggtcgg gctgaacggg gggttcgtgc acacagccca
     5461 gcttggagcg aacgacctac accgaactga gatacctaca gcgtgagcta tgagaaagcg
     5521 ccacgcttcc cgaagggaga aaggcggaca ggtatccggt aagcggcagg gtcggaacag
     5581 gagagcgcac gagggagctt ccagggggaa acgcctggta tctttatagt cctgtcgggt
     5641 ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc gtcagggggg cggagcctat
     5701 ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc cttttgctgg ccttttgctc
     5761 acatgtcctg caggcagctg c
//
