LOCUS       sequence-333908-        6214 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Encodes Cre-dependent procaspase 3 and TEVp under control of the 
            TRE.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6214)
  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 6214)
  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-TRE-DIO-proCaspase-3-TEVp
FEATURES             Location/Qualifiers
     source          1..6214
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     protein_bind    40..58
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    82..100
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    124..142
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    166..184
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    208..226
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     repeat_region   383..523
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   383..512
                     /label=AAV2 ITR
     rep_origin      598..1053
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(685..704)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     895..916
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1070..1089)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     1189..1211
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(1249..1267)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        1335..1439
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1440..1508,1509..2300)
                     /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(1658..1677)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2471..3059
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2960..2979
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   3121..3250
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     polyA_signal    3273..3480
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     3469..3486
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     misc_feature    3521..4109
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             3627..3638
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     4036..4056
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    complement(4133..4166)
                     /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(4177..4210)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             4768..4788
                     /codon_start=1
                     /product="tobacco etch virus (TEV) protease recognition and
                     cleavage site"
                     /label=TEV site
                     /translation="ENLYFQS"
     CDS             5104..5157
                     /codon_start=1
                     /product="2A peptide from Thosea asigna virus capsid 
                     protein"
                     /label=T2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="EGRGSLLTCGDVEENPGP"
     protein_bind    5929..5962
                     /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    5973..6006
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     promoter        complement(6141..6179)
                     /label=minimal CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     protein_bind    join(6212..6214,1..16)
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
ORIGIN
        1 ctatcactga tagggagtgg taaactcgac tttcactttt ctctatcact gatagggagt
       61 ggtaaactcg actttcactt ttctctatca ctgataggga gtggtaaact cgactttcac
      121 ttttctctat cactgatagg gagtggtaaa ctcgactttc acttttctct atcactgata
      181 gggagtggta aactcgactt tcacttttct ctatcactga tagggagtgg taaactcgac
      241 ctatataagc agagctcgtt tagtgaaccg tcagatcgcc tggagacgcc atccacgctg
      301 ttttgacctc catagaagac accgggaccg atccagcctc cgcggccccg aattcctgca
      361 gaattgtttc gcgtgcggcc gcaggaaccc ctagtgatgg agttggccac tccctctctg
      421 cgcgctcgct cgctcactga ggccgggcga ccaaaggtcg cccgacgccc gggctttgcc
      481 cgggcggcct cagtgagcga gcgagcgcgc agctgcctgc aggggcgcct gatgcggtat
      541 tttctcctta cgcatctgtg cggtatttca caccgcatac gtcaaagcaa ccatagtacg
      601 cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt tacgcgcagc gtgaccgcta
      661 cacttgccag cgccttagcg cccgctcctt tcgctttctt cccttccttt ctcgccacgt
      721 tcgccggctt tccccgtcaa gctctaaatc gggggctccc tttagggttc cgatttagtg
      781 ctttacggca cctcgacccc aaaaaacttg atttgggtga tggttcacgt agtgggccat
      841 cgccctgata gacggttttt cgccctttga cgttggagtc cacgttcttt aatagtggac
      901 tcttgttcca aactggaaca acactcaact ctatctcggg ctattctttt gatttataag
      961 ggattttgcc gatttcggtc tattggttaa aaaatgagct gatttaacaa aaatttaacg
     1021 cgaattttaa caaaatatta acgtttacaa ttttatggtg cactctcagt acaatctgct
     1081 ctgatgccgc atagttaagc cagccccgac acccgccaac acccgctgac gcgccctgac
     1141 gggcttgtct gctcccggca tccgcttaca gacaagctgt gaccgtctcc gggagctgca
     1201 tgtgtcagag gttttcaccg tcatcaccga aacgcgcgag acgaaagggc ctcgtgatac
     1261 gcctattttt ataggttaat gtcatgataa taatggtttc ttagacgtca ggtggcactt
     1321 ttcggggaaa tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt
     1381 atccgctcat gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta
     1441 tgagtattca acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg
     1501 tttttgctca cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac
     1561 gagtgggtta catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg
     1621 aagaacgttt tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc
     1681 gtattgacgc cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg
     1741 ttgagtactc accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat
     1801 gcagtgctgc cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg
     1861 gaggaccgaa ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg
     1921 atcgttggga accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc
     1981 ctgtagcaat ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt
     2041 cccggcaaca attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct
     2101 cggcccttcc ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc
     2161 gcggtatcat tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca
     2221 cgacggggag tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct
     2281 cactgattaa gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt
     2341 taaaacttca tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga
     2401 ccaaaatccc ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca
     2461 aaggatcttc ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac
     2521 caccgctacc agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg
     2581 taactggctt cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag
     2641 gccaccactt caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac
     2701 cagtggctgc tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt
     2761 taccggataa ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg
     2821 agcgaacgac ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc
     2881 ttcccgaagg gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc
     2941 gcacgaggga gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc
     3001 acctctgact tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa
     3061 acgccagcaa cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt
     3121 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
     3181 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
     3241 aggggttcct gcggccgctc ggtccgcacg tgtccccagc atgcctgcta ttgtcttccc
     3301 aatcctcccc cttgctgtcc tgccccaccc caccccccag aatagaatga cacctactca
     3361 gacaatgcga tgcaatttcc tcattttatt aggaaaggac agtgggagtg gcaccttcca
     3421 gggtcaagga aggcacgggg gaggggcaaa caacagatgg ctggcaacta gaaggcacag
     3481 tcgaggcaga tcgatctctc gagcagcgct cggtatcgat gcggggaggc ggcccaaagg
     3541 gagatccgac tcgtctgagg gcgaaggcga agacgcggaa gaggccgcag agccggcagc
     3601 aggccgcggg aaggaaggtc cgctggattg agggccgaag ggacgtagca gaaggacgtc
     3661 ccgcgcagaa tccaggtggc aacataggcg agcagccaag gaaaggacga tgatttcccc
     3721 gacaacacca cggaattgtc agtgcccaac agccgagccc ctgtccagca gcgggcaagg
     3781 caggcggcga tgagttccgc cgtggcaata gggaggggga aagcgaaagt cccggaaagg
     3841 agctgacagg tggtggcaat gccccaacca gtgggggttg cgtcagcaaa cacagtgcac
     3901 accacgccac gttgcctgac aacgggccac aactcctcat aaagagacag caaccaggat
     3961 ttatacaagg aggagaaaat gaaagccata cgggaagcaa tagcatgata caaaggcatt
     4021 aaagcagcgt atccacatag cgtaaaagga gcaacatagt taagaatacc agtcaatctt
     4081 tcacaaattt tgtaatccag aggttgatta tcgataagct tgatgatctt cgataacttc
     4141 gtataggata ctttatacga agttattgcg attcggataa cttcgtatag catacattat
     4201 acgaagttat caaggccggc caggttggcg cgccaactag ctagcatgga gaacactgaa
     4261 aactcagtgg attcaaaatc cattaaaaat ttggaaccaa agatcataca tggaagcgaa
     4321 tcaatggact ctggaatatc cctggacaac agttataaaa tggattatcc tgagatgggt
     4381 ttatgtataa taattaataa taagaatttt cataaaagca ctggaatgac atctcggtct
     4441 ggtacagatg tcgatgcagc aaacctcagg gaaacattca gaaacttgaa atatgaagtc
     4501 aggaataaaa atgatcttac acgtgaagaa attgtggaat tgatgcgtga tgtttctaaa
     4561 gaagatcaca gcaaaaggag cagttttgtt tgtgtgcttc tgagccatgg tgaagaagga
     4621 ataatttttg gaacaaatgg acctgttgac ctgaaaaaaa taacaaactt tttcagaggg
     4681 gatcgttgta gaagtctaac tggaaaaccc aaacttttca ttattcaggc ctgccgtggt
     4741 acagaactgg actgtggcat tgagacagaa aatctctact tccagagtgg tgttgatgat
     4801 gacatggcgt gtcataaaat accagtggag gccgacttct tgtatgcata ctccacagca
     4861 cctggttatt attcttggcg aaattcaaag gatggctcct ggttcatcca gtcgctttgt
     4921 gccatgctga aacagtatgc cgacaagctt gaatttatgc acattcttac ccgggttaac
     4981 cgaaaggtgg caacagaatt tgagtccttt tcctttgacg ctacttttca tgcaaagaaa
     5041 cagattccat gtattgtttc catgctcaca aaagaactct atttttatca ctccggaagc
     5101 ggagagggca gaggaagtct tctaacatgc ggtgacgtgg aggagaatcc cggccctggc
     5161 gagagccttt tcaagggccc gagggactac aacccgatct ccagcaccat ctgtcacctg
     5221 accaacgaga gcgacggtca caccactagt ctgtacggca tcggcttcgg ccccttcatc
     5281 atcaccaaca agcatctgtt caggaggaat aacggcacac tgctggtgca aagcctgcac
     5341 ggcgtgttca aagtgaagaa cacaaccacc ctgcaacagc acctgatcga cggcagggac
     5401 atgattatca tcaggatgcc caaggacttc cccccctttc cccagaaact gaagttcagg
     5461 gagccacaaa gggaggagcg aatctgcctg gtgaccacca acttccagac caagtccatg
     5521 agcagcatgg tctctgatac cagctgcacc ttccccagca gcgacggcat cttctggaag
     5581 cactggattc agacgaagga tggccaatgc ggcagcccat tggtgagcac tagggacggc
     5641 ttcatcgtgg gcatccacag cgccagcaat tttaccaata ccaacaacta cttcacgagc
     5701 gtgccgaaaa acttcatgga gctgttgacc aatcaagagg cgcagcagtg ggtgagcggc
     5761 tggaggctga acgccgacag cgttctttgg ggcggacata aggtgttcat ggtcaagccc
     5821 gaggaaccct tccagcccgt taaggaagcc actcagcttt gatgtacaag taagaattcc
     5881 ttgggccccc tcgaaggtac gttgcgttat ttaaatttgg cgcgccaaat aacttcgtat
     5941 aaagtatcct atacgaagtt attgcgattc ggataacttc gtataatgta tgctatacga
     6001 agttatcgac gtcgactaga ggatccccgg gtaccgagct cgaattcggg gccgcggagg
     6061 ctggatcggt cccggtgtct tctatggagg tcaaaacagc gtggatggcg tctccaggcg
     6121 atctgacggt tcactaaacg agctctgctt atataggcct cccaccgtac acgcctacct
     6181 cgacccgggt accgagctcg actttcactt ttct
//
