LOCUS       sequence-178870-        7779 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  DO/DIO Cre-Switch. Expresses tdTomato in Cre negative cells, 
            expresses EGFP in Cre positive cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7779)
  AUTHORS   Saunders A, Johnson CA, Sabatini BL
  TITLE     Novel recombinant adeno-associated viruses for Cre activated and 
            inactivated transgene expression in neurons.
  JOURNAL   Front Neural Circuits. 2012 Jul 27;6:47. doi: 
            10.3389/fncir.2012.00047. eCollection 2012.
  PUBMED    22866029
REFERENCE   2  (bases 1 to 7779)
  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: "Front 
            Neural Circuits."; date: "2012-07-27"; volume: "6:47. doi"; pages: "
            10.3389/fncir.2012.00047. eCollection 2012"
COMMENT     Sequence Label: pAAV-Ef1a-DO_DIO-TdTomato_EGFP-WPRE-pA
FEATURES             Location/Qualifiers
     source          1..7779
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..130
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        231..1409
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          462..1400
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1357..1377
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1440..1473)
                     /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(1524..1557)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      1579..1588
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1585..3015
                     /codon_start=1
                     /product="tandem dimeric (pseudo-monomeric) derivative of 
                     DsRed (Shaner et al., 2004)"
                     /label=tdTomato
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEEVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPYEGTQTA
                     KLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVMNFEDGG
                     LVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGVLKGEIH
                     QALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYERSEGRH
                     HLFLGHGTGSTGSGSSGTASSEDNNMAVIKEFMRFKVRMEGSMNGHEFEIEGEGEGRPY
                     EGTQTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYKKLSFPEGFKWERVM
                     NFEDGGLVTVTQDSSLQDGTLIYKVKMRGTNFPPDGPVMQKKTMGWEASTERLYPRDGV
                     LKGEIHQALKLKDGGHYLVEFKTIYMAKKPVQLPGYYYVDTKLDITSHNEDYTIVEQYE
                     RSEGRHHLFLYGMDELYK"
     CDS             complement(join(3016..3729,3730..3732,3733..3735))
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     CDS             complement(join(3019..3729,3730..3732,3733..3735))
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="MVSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTL
                     KFICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDD
                     GNYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIK
                     VNFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLL
                     EFVTAAGITLGMDELYK"
     primer_bind     complement(3061..3082)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     3410..3429
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3669..3690
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     regulatory      3732..3741
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3764..3797
                     /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    3848..3881
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3906..4494
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3959..3979)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4377..4388)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4526..5002
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4725..4744)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   5042..5182
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   5042..5171
                     /label=AAV2 ITR
     rep_origin      5257..5712
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5344..5363)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5554..5575
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5729..5748)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5848..5870
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5908..5926)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5994..6098
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6099..6167,6168..6959)
                     /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(6317..6336)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7130..7718
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7619..7638
                     /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 gcgtaagctt tgcaaagatg gataaagttt taaacagaga
      181 ggaatctttg cagctaatgg accttctagg tcttgaaagg agtgggaatt ggctccggtg
      241 cccgtcagtg ggcagagcgc acatcgccca cagtccccga gaagttgggg ggaggggtcg
      301 gcaattgaac cggtgcctag agaaggtggc gcggggtaaa ctgggaaagt gatgtcgtgt
      361 actggctccg cctttttccc gagggtgggg gagaaccgta tataagtgca gtagtcgccg
      421 tgaacgttct ttttcgcaac gggtttgccg ccagaacaca ggtaagtgcc gtgtgtggtt
      481 cccgcgggcc tggcctcttt acgggttatg gcccttgcgt gccttgaatt acttccacct
      541 ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg gagagttcga
      601 ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg cctgggcgct
      661 ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct ttcgataagt
      721 ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg caagatagtc
      781 ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc gcgggcggcg
      841 acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga gcgcggccac
      901 cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct ggcctcgcgc
      961 cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca gttgcgtgag
     1021 cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg acgcggcgct
     1081 cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg tcctcagccg
     1141 tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat tagttctcga
     1201 gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg gagtttcccc
     1261 acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa ttctccttgg
     1321 aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca gtggttcaaa
     1381 gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagtc gactccggaa
     1441 taacttcgta taggatactt tatacgaagt tatgcagaat ggtagctgga ttgtagctgc
     1501 tattagcaat atgaaacctc ttaataactt cgtatagcat acattatacg aagttatggc
     1561 gcgccttatc tagatttagc caccatggtg agcaagggcg aggaggtcat caaagagttc
     1621 atgcgcttca aggtgcgcat ggagggctcc atgaacggcc acgagttcga gatcgagggc
     1681 gagggcgagg gccgccccta cgagggcacc cagaccgcca agctgaaggt gaccaagggc
     1741 ggccccctgc ccttcgcctg ggacatcctg tccccccagt tcatgtacgg ctccaaggcg
     1801 tacgtgaagc accccgccga catccccgat tacaagaagc tgtccttccc cgagggcttc
     1861 aagtgggagc gcgtgatgaa cttcgaggac ggcggtctgg tgaccgtgac ccaggactcc
     1921 tccctgcagg acggcacgct gatctacaag gtgaagatgc gcggcaccaa cttccccccc
     1981 gacggccccg taatgcagaa gaagaccatg ggctgggagg cctccaccga gcgcctgtac
     2041 ccccgcgacg gcgtgctgaa gggcgagatc caccaggccc tgaagctgaa ggacggcggc
     2101 cactacctgg tggagttcaa gaccatctac atggccaaga agcccgtgca actgcccggc
     2161 tactactacg tggacaccaa gctggacatc acctcccaca acgaggacta caccatcgtg
     2221 gaacagtacg agcgctccga gggccgccac cacctgttcc tggggcatgg caccggcagc
     2281 accggcagcg gcagctccgg caccgcctcc tccgaggaca acaacatggc cgtcatcaaa
     2341 gagttcatgc gcttcaaggt gcgcatggag ggctccatga acggccacga gttcgagatc
     2401 gagggcgagg gcgagggccg cccctacgag ggcacccaga ccgccaagct gaaggtgacc
     2461 aagggcggcc ccctgccctt cgcctgggac atcctgtccc cccagttcat gtacggctcc
     2521 aaggcgtacg tgaagcaccc cgccgacatc cccgattaca agaagctgtc cttccccgag
     2581 ggcttcaagt gggagcgcgt gatgaacttc gaggacggcg gtctggtgac cgtgacccag
     2641 gactcctccc tgcaggacgg cacgctgatc tacaaggtga agatgcgcgg caccaacttc
     2701 ccccccgacg gccccgtaat gcagaagaag accatgggct gggaggcctc caccgagcgc
     2761 ctgtaccccc gcgacggcgt gctgaagggc gagatccacc aggccctgaa gctgaaggac
     2821 ggcggccact acctggtgga gttcaagacc atctacatgg ccaagaagcc cgtgcaactg
     2881 cccggctact actacgtgga caccaagctg gacatcacct cccacaacga ggactacacc
     2941 atcgtggaac agtacgagcg ctccgagggc cgccaccacc tgttcctgta cggcatggac
     3001 gagctgtaca agtaattact tgtacagctc gtccatgccg agagtgatcc cggcggcggt
     3061 cacgaactcc agcaggacca tgtgatcgcg cttctcgttg gggtctttgc tcagggcgga
     3121 ctgggtgctc aggtagtggt tgtcgggcag cagcacgggg ccgtcgccga tgggggtgtt
     3181 ctgctggtag tggtcggcga gctgcacgct gccgtcctcg atgttgtggc ggatcttgaa
     3241 gttcaccttg atgccgttct tctgcttgtc ggccatgata tagacgttgt ggctgttgta
     3301 gttgtactcc agcttgtgcc ccaggatgtt gccgtcctcc ttgaagtcga tgcccttcag
     3361 ctcgatgcgg ttcaccaggg tgtcgccctc gaacttcacc tcggcgcggg tcttgtagtt
     3421 gccgtcgtcc ttgaagaaga tggtgcgctc ctggacgtag ccttcgggca tggcggactt
     3481 gaagaagtcg tgctgcttca tgtggtcggg gtagcggctg aagcactgca cgccgtaggt
     3541 cagggtggtc acgagggtgg gccagggcac gggcagcttg ccggtggtgc agatgaactt
     3601 cagggtcagc ttgccgtagg tggcatcgcc ctcgccctcg ccggacacgc tgaacttgtg
     3661 gccgtttacg tcgccgtcca gctcgaccag gatgggcacc accccggtga acagctcctc
     3721 gcccttgctc accatggtgg cggtggcgac cggtagcgct agcataactt cgtataaagt
     3781 atcctatacg aagttatttg ccttaaccca gaaattatca ctgttattct ttagaatggt
     3841 gcaaagaata acttcgtata atgtatgcta tacgaagtta tgaattcgat atcaagctta
     3901 tcgataatca acctctggat tacaaaattt gtgaaagatt gactggtatt cttaactatg
     3961 ttgctccttt tacgctatgt ggatacgctg ctttaatgcc tttgtatcat gctattgctt
     4021 cccgtatggc tttcattttc tcctccttgt ataaatcctg gttgctgtct ctttatgagg
     4081 agttgtggcc cgttgtcagg caacgtggcg tggtgtgcac tgtgtttgct gacgcaaccc
     4141 ccactggttg gggcattgcc accacctgtc agctcctttc cgggactttc gctttccccc
     4201 tccctattgc cacggcggaa ctcatcgccg cctgccttgc ccgctgctgg acaggggctc
     4261 ggctgttggg cactgacaat tccgtggtgt tgtcggggaa atcatcgtcc tttccttggc
     4321 tgctcgccta tgttgccacc tggattctgc gcgggacgtc cttctgctac gtcccttcgg
     4381 ccctcaatcc agcggacctt ccttcccgcg gcctgctgcc ggctctgcgg cctcttccgc
     4441 gtcttcgcct tcgccctcag acgagtcgga tctccctttg ggccgcctcc ccgcatcgat
     4501 accgagcgct gctcgagaga tctacgggtg gcatccctgt gacccctccc cagtgcctct
     4561 cctggccctg gaagttgcca ctccagtgcc caccagcctt gtcctaataa aattaagttg
     4621 catcattttg tctgactagg tgtccttcta taatattatg gggtggaggg gggtggtatg
     4681 gagcaagggg caagttggga agacaacctg tagggcctgc ggggtctatt gggaaccaag
     4741 ctggagtgca gtggcacaat cttggctcac tgcaatctcc gcctcctggg ttcaagcgat
     4801 tctcctgcct cagcctcccg agttgttggg attccaggca tgcatgacca ggctcagcta
     4861 atttttgttt ttttggtaga gacggggttt caccatattg gccaggctgg tctccaactc
     4921 ctaatctcag gtgatctacc caccttggcc tcccaaattg ctgggattac aggcgtgaac
     4981 cactgctccc ttccctgtcc ttctgatttt gtaggtaacc acgtgcggac cgagcggccg
     5041 caggaacccc tagtgatgga gttggccact ccctctctgc gcgctcgctc gctcactgag
     5101 gccgggcgac caaaggtcgc ccgacgcccg ggctttgccc gggcggcctc agtgagcgag
     5161 cgagcgcgca gctgcctgca ggggcgcctg atgcggtatt ttctccttac gcatctgtgc
     5221 ggtatttcac accgcatacg tcaaagcaac catagtacgc gccctgtagc ggcgcattaa
     5281 gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac acttgccagc gccttagcgc
     5341 ccgctccttt cgctttcttc ccttcctttc tcgccacgtt cgccggcttt ccccgtcaag
     5401 ctctaaatcg ggggctccct ttagggttcc gatttagtgc tttacggcac ctcgacccca
     5461 aaaaacttga tttgggtgat ggttcacgta gtgggccatc gccctgatag acggtttttc
     5521 gccctttgac gttggagtcc acgttcttta atagtggact cttgttccaa actggaacaa
     5581 cactcaactc tatctcgggc tattcttttg atttataagg gattttgccg atttcggtct
     5641 attggttaaa aaatgagctg atttaacaaa aatttaacgc gaattttaac aaaatattaa
     5701 cgtttacaat tttatggtgc actctcagta caatctgctc tgatgccgca tagttaagcc
     5761 agccccgaca cccgccaaca cccgctgacg cgccctgacg ggcttgtctg ctcccggcat
     5821 ccgcttacag acaagctgtg accgtctccg ggagctgcat gtgtcagagg ttttcaccgt
     5881 catcaccgaa acgcgcgaga cgaaagggcc tcgtgatacg cctattttta taggttaatg
     5941 tcatgataat aatggtttct tagacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     6001 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     6061 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     6121 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     6181 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     6241 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     6301 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     6361 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     6421 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     6481 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     6541 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     6601 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     6661 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     6721 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     6781 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     6841 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     6901 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     6961 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     7021 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     7081 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     7141 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     7201 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     7261 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     7321 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     7381 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     7441 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     7501 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     7561 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     7621 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     7681 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     7741 tacggttcct ggccttttgc tggccttttg ctcacatgt
//
