LOCUS       sequence-190883-        5454 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  An AAV genome with tet-inducible, Cre-dependent expression of the 
            fluorescent protein mRuby2.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5454)
  AUTHORS   Chan KY, Jang MJ, Yoo BB, Greenbaum A, Ravi N, Wu WL, 
            Sanchez-Guardado L, Lois C, Mazmanian SK, Deverman BE, Gradinaru V
  TITLE     Engineered AAVs for efficient noninvasive gene delivery to the 
            central and peripheral nervous systems.
  JOURNAL   Nat Neurosci. 2017 Aug;20(8):1172-1179. doi: 10.1038/nn.4593. Epub 
            2017 Jun 26.
  PUBMED    28671695
REFERENCE   2  (bases 1 to 5454)
  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"; doi: "10.1038/nn.4593"; 
            journalName: "Nat Neurosci"; date: "2017-08"; volume: "20"; issue: 
            "8"; pages: "1172-1179"
COMMENT     Sequence Label: pAAV-TRE-DIO-mRuby2
FEATURES             Location/Qualifiers
     source          1..5454
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     protein_bind    165..183
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    202..220
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    239..257
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    276..294
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    313..331
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    350..368
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    387..405
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     protein_bind    424..442
                     /gene="tetO"
                     /label=tet operator
                     /bound_moiety="tetracycline repressor TetR"
                     /note="bacterial operator O2 for the tetR and tetA genes"
     primer_bind     464..488
                     /label=LNCX
                     /note="Human CMV promoter, forward primer"
     protein_bind    complement(587..620)
                     /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(671..704)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(716..1426)
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     regulatory      1423..1432
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    1439..1472
                     /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    1523..1556
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    1581..2169
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(1634..1654)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(2052..2063)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    2201..2677
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(2400..2419)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   2717..2857
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   2717..2846
                     /label=AAV2 ITR
     rep_origin      2932..3387
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3019..3038)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3229..3250
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3404..3423)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     3523..3545
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(3583..3601)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        3669..3773
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3774..3842,3843..4634)
                     /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(3992..4011)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4805..5393
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5294..5313
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcgtcg ggcgaccttt
       61 ggtcgcccgg cctcagtgag cgagcgagcg cgcagagagg gagtggccaa ctccatcact
      121 aggggttcct gcggccgcac gcgtatcaag tgccacctga cgtctcccta tcagtgatag
      181 agaagtcgac acgtctcgag ctccctatca gtgatagaga aggtacgtct agaacgtctc
      241 cctatcagtg atagagaagt cgacacgtct cgagctccct atcagtgata gagaaggtac
      301 gtctagaacg tctccctatc agtgatagag aagtcgacac gtctcgagct ccctatcagt
      361 gatagagaag gtacgtctag aacgtctccc tatcagtgat agagaagtcg acacgtctcg
      421 agctccctat cagtgataga gaagctaccc cctatataag cagagctcgt ttagtgaacc
      481 gtcagatcgc ctggagacgc catccacgct gttttgacct ccatagaaga caccgggacc
      541 gatccagcct ggatccggta ccggatcctc tagagtcgac tccggaataa cttcgtatag
      601 gatactttat acgaagttat gcagaatggt agctggattg tagctgctat tagcaatatg
      661 aaacctctta ataacttcgt atagcataca ttatacgaag ttatggcgcg cctcatttgt
      721 acagctcatc cattcctccg ccaaggccag caaacttggc aactgcatgt tcccgttgta
      781 ccacaaacat ctcgttgtca gactcctcca gtcgttccaa gcggtgatca accgcatgta
      841 tgccaggcat tttgatgttg ccgacagttt tcttgctcct gtacgtggtc acaaatgagc
      901 atgacaagtg gcctcctccg tcaaccttca gagccatgtg cgtatagcct ctcagtcctc
      961 catctgcagg atacatcatc tctgtgtttg gctcccaccc tttggttttc ttctgcatca
     1021 caggcccatt ggagggaaag ttcacccctc tcacttgcac gtgatagacc agacatccgt
     1081 cctccagact agtgtcctgc atgactgtga ccacaccacc atcctcgtaa cgggtcaccc
     1141 tttcccatgt gaagccctct ggaaaggact gtttgaagaa gtctggaatg cctttggggt
     1201 acttgatgaa ggtccgactg ccatacatga agctagtggc aagaatgtcg aaggcaaacg
     1261 ggagagggcc accttcaata accttgatcc tcatggtctg agtacccatg tagggattcc
     1321 cttcaccttc ccctgtgcac ttgaactgat gcccattcac agacccttcc atgactacct
     1381 tcatgcgcat gttctccttg atgagttcct ctcccttgct caccatggtg gcgctagcat
     1441 aacttcgtat aaagtatcct atacgaagtt atttgcctta acccagaaat tatcactgtt
     1501 attctttaga atggtgcaaa gaataacttc gtataatgta tgctatacga agttatgaat
     1561 tcgatatcaa gcttatcgat aatcaacctc tggattacaa aatttgtgaa agattgactg
     1621 gtattcttaa ctatgttgct ccttttacgc tatgtggata cgctgcttta atgcctttgt
     1681 atcatgctat tgcttcccgt atggctttca ttttctcctc cttgtataaa tcctggttgc
     1741 tgtctcttta tgaggagttg tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt
     1801 ttgctgacgc aacccccact ggttggggca ttgccaccac ctgtcagctc ctttccggga
     1861 ctttcgcttt ccccctccct attgccacgg cggaactcat cgccgcctgc cttgcccgct
     1921 gctggacagg ggctcggctg ttgggcactg acaattccgt ggtgttgtcg gggaaatcat
     1981 cgtcctttcc ttggctgctc gcctatgttg ccacctggat tctgcgcggg acgtccttct
     2041 gctacgtccc ttcggccctc aatccagcgg accttccttc ccgcggcctg ctgccggctc
     2101 tgcggcctct tccgcgtctt cgccttcgcc ctcagacgag tcggatctcc ctttgggccg
     2161 cctccccgca tcgataccga gcgctgctcg agagatctac gggtggcatc cctgtgaccc
     2221 ctccccagtg cctctcctgg ccctggaagt tgccactcca gtgcccacca gccttgtcct
     2281 aataaaatta agttgcatca ttttgtctga ctaggtgtcc ttctataata ttatggggtg
     2341 gaggggggtg gtatggagca aggggcaagt tgggaagaca acctgtaggg cctgcggggt
     2401 ctattgggaa ccaagctgga gtgcagtggc acaatcttgg ctcactgcaa tctccgcctc
     2461 ctgggttcaa gcgattctcc tgcctcagcc tcccgagttg ttgggattcc aggcatgcat
     2521 gaccaggctc agctaatttt tgtttttttg gtagagacgg ggtttcacca tattggccag
     2581 gctggtctcc aactcctaat ctcaggtgat ctacccacct tggcctccca aattgctggg
     2641 attacaggcg tgaaccactg ctcccttccc tgtccttctg attttgtagg taaccacgtg
     2701 cggaccgagc ggccgcagga acccctagtg atggagttgg ccactccctc tctgcgcgct
     2761 cgctcgctca ctgaggccgg gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg
     2821 gcctcagtga gcgagcgagc gcgcagctgc ctgcaggggc gcctgatgcg gtattttctc
     2881 cttacgcatc tgtgcggtat ttcacaccgc atacgtcaaa gcaaccatag tacgcgccct
     2941 gtagcggcgc attaagcgcg gcgggtgtgg tggttacgcg cagcgtgacc gctacacttg
     3001 ccagcgcctt agcgcccgct cctttcgctt tcttcccttc ctttctcgcc acgttcgccg
     3061 gctttccccg tcaagctcta aatcgggggc tccctttagg gttccgattt agtgctttac
     3121 ggcacctcga ccccaaaaaa cttgatttgg gtgatggttc acgtagtggg ccatcgccct
     3181 gatagacggt ttttcgccct ttgacgttgg agtccacgtt ctttaatagt ggactcttgt
     3241 tccaaactgg aacaacactc aactctatct cgggctattc ttttgattta taagggattt
     3301 tgccgatttc ggtctattgg ttaaaaaatg agctgattta acaaaaattt aacgcgaatt
     3361 ttaacaaaat attaacgttt acaattttat ggtgcactct cagtacaatc tgctctgatg
     3421 ccgcatagtt aagccagccc cgacacccgc caacacccgc tgacgcgccc tgacgggctt
     3481 gtctgctccc ggcatccgct tacagacaag ctgtgaccgt ctccgggagc tgcatgtgtc
     3541 agaggttttc accgtcatca ccgaaacgcg cgagacgaaa gggcctcgtg atacgcctat
     3601 ttttataggt taatgtcatg ataataatgg tttcttagac gtcaggtggc acttttcggg
     3661 gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat atgtatccgc
     3721 tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag agtatgagta
     3781 ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt cctgtttttg
     3841 ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt gcacgagtgg
     3901 gttacatcga actggatctc aacagcggta agatccttga gagttttcgc cccgaagaac
     3961 gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta tcccgtattg
     4021 acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac ttggttgagt
     4081 actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa ttatgcagtg
     4141 ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg atcggaggac
     4201 cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc cttgatcgtt
     4261 gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg atgcctgtag
     4321 caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta gcttcccggc
     4381 aacaattaat agactggatg gaggcggata aagttgcagg accacttctg cgctcggccc
     4441 ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg tctcgcggta
     4501 tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc tacacgacgg
     4561 ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt gcctcactga
     4621 ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt gatttaaaac
     4681 ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc atgaccaaaa
     4741 tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag atcaaaggat
     4801 cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa aaaccaccgc
     4861 taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg aaggtaactg
     4921 gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag ttaggccacc
     4981 acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg ttaccagtgg
     5041 ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga tagttaccgg
     5101 ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc ttggagcgaa
     5161 cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc acgcttcccg
     5221 aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga gagcgcacga
     5281 gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt cgccacctct
     5341 gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg aaaaacgcca
     5401 gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac atgt
//
