LOCUS       sequence-397599-        7045 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Double floxed red fluorescent protein tdTomato in AAV production 
            vector expressed under the mammalian promoter (EF1a).
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7045)
  AUTHORS   Lu X, Wang Y, Liu Z, Gou Y, Jaeger D, St-Pierre F
  TITLE     Widefield imaging of rapid pan-cortical voltage dynamics with an 
            indicator evolved for one-photon microscopy.
  JOURNAL   Nat Commun. 2023 Oct 12;14(1):6423. doi: 10.1038/s41467-023-41975-3.
  PUBMED    37828037
REFERENCE   2  (bases 1 to 7045)
  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: "Nat 
            Commun."; date: "2023-10-12"; volume: "14(1):6423. doi"; pages: " 
            10.1038/s41467-023-41975-3"
COMMENT     Sequence Label: pAAV-EF1a-DIO-tdTomato-WPRE
FEATURES             Location/Qualifiers
     source          1..7045
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   2..131
                     /label=AAV2 ITR (alternate)
                     /note="Functional equivalent of wild-type AAV2 ITR"
     promoter        232..1410
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          463..1401
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1358..1378
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    complement(1441..1474)
                     /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(1525..1558)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     regulatory      3011..3020
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3031..3064
                     /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    3115..3148
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3173..3761
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3226..3246)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3644..3655)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3793..4269
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3992..4011)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4309..4449
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4309..4438
                     /label=AAV2 ITR
     rep_origin      4524..4979
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4611..4630)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4821..4842
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4996..5015)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5115..5137
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5175..5193)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5261..5365
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5366..5434,5435..6226)
                     /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(5584..5603)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6397..6985
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6886..6905
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 tcctgcaggc agctgcgcgc tcgctcgctc actgaggccg cccgggcgtc gggcgacctt
       61 tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca actccatcac
      121 taggggttcc tgcggccgca cgcgtaagct ttgcaaagat ggataaagtt ttaaacagag
      181 aggaatcttt gcagctaatg gaccttctag gtcttgaaag gagtgggaat tggctccggt
      241 gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg gggaggggtc
      301 ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag tgatgtcgtg
      361 tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc agtagtcgcc
      421 gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc cgtgtgtggt
      481 tcccgcgggc ctggcctctt tacgggttat ggcccttgcg tgccttgaat tacttccacc
      541 tggctgcagt acgtgattct tgatcccgag cttcgggttg gaagtgggtg ggagagttcg
      601 aggccttgcg cttaaggagc cccttcgcct cgtgcttgag ttgaggcctg gcctgggcgc
      661 tggggccgcc gcgtgcgaat ctggtggcac cttcgcgcct gtctcgctgc tttcgataag
      721 tctctagcca tttaaaattt ttgatgacct gctgcgacgc tttttttctg gcaagatagt
      781 cttgtaaatg cgggccaaga tctgcacact ggtatttcgg tttttggggc cgcgggcggc
      841 gacggggccc gtgcgtccca gcgcacatgt tcggcgaggc ggggcctgcg agcgcggcca
      901 ccgagaatcg gacgggggta gtctcaagct ggccggcctg ctctggtgcc tggcctcgcg
      961 ccgccgtgta tcgccccgcc ctgggcggca aggctggccc ggtcggcacc agttgcgtga
     1021 gcggaaagat ggccgcttcc cggccctgct gcagggagct caaaatggag gacgcggcgc
     1081 tcgggagagc gggcgggtga gtcacccaca caaaggaaaa gggcctttcc gtcctcagcc
     1141 gtcgcttcat gtgactccac ggagtaccgg gcgccgtcca ggcacctcga ttagttctcg
     1201 agcttttgga gtacgtcgtc tttaggttgg ggggaggggt tttatgcgat ggagtttccc
     1261 cacactgagt gggtggagac tgaagttagg ccagcttggc acttgatgta attctccttg
     1321 gaatttgccc tttttgagtt tggatcttgg ttcattctca agcctcagac agtggttcaa
     1381 agtttttttc ttccatttca ggtgtcgtga ggtaccggat cctctagagt cgactccgga
     1441 ataacttcgt ataggatact ttatacgaag ttatgcagaa tggtagctgg attgtagctg
     1501 ctattagcaa tatgaaacct cttaataact tcgtatagca tacattatac gaagttatgg
     1561 cgcgccctat tacttgtaca tcattactta tacagctcat ccatgccgta cagaaacaga
     1621 tggtggcgtc cttcagacct ctcatactgt tccacaatgg tgtagtcttc attgtgggat
     1681 gtgatatcca gcttggtgtc cacatagtaa tagccgggca gctgcactgg tttcttggcc
     1741 atgtagatgg ttttgaactc caccaggtag tggccgccgt ctttcagctt cagggcctgg
     1801 tggatttcgc ccttcagcac tccgtctcta ggatacagcc gttcagtgct ggcctcccac
     1861 cccattgttt tcttctgcat cacgggccca tcgggtggaa aattggtgcc gcgcattttg
     1921 actttgtaga tcagagtgcc atcctgcaga gaggagtcct gtgtcacggt caccaggcct
     1981 ccatcttcaa aattcatcac tctctcccac ttaaagccct ctgggaaaga cagtttcttg
     2041 taatcaggaa tgtcagcggg atgcttgaca taggccttac ttccgtacat aaactgagga
     2101 gacaggatat cccaagcgaa aggcagtggt cctcccttgg tcactttcag cttggcagtc
     2161 tgtgtgccct cgtaaggcct cccttcgccc tctccttcaa tctcaaattc gtgcccattc
     2221 atgctgccct ccatgcgcac cttaaacctc atgaactctt tgatcacagc catattgtta
     2281 tcctcgcttg aggcggttcc actagaccct gagccggtac ttccagtccc atggcccagg
     2341 aacagatggt gtctcccctc gctccgttca tactgctcca caatagtgta gtcctcgttg
     2401 tgtgaagtga tatccagttt tgtgtccacg taatagtagc ctggcagctg cacaggtttc
     2461 ttggccatgt agattgtctt gaactccacc aggtaatgtc ccccatcttt cagcttcaga
     2521 gcctggtgga tttctccttt cagcaccccg tcgcgtggat acagcctttc ggtggaggcc
     2581 tcccatccca tagttttctt ctgcatcaca ggtccatcag ggggaaaatt tgtccctctc
     2641 attttcacct tgtagatcag ggtcccatcc tgcagggagc tgtcctgggt cacagtcacc
     2701 aggcctccgt cctcgaagtt catcactctt tcccacttaa acccctcggg gaagctcagt
     2761 ttcttataat cgggaatgtc agctggatgt ttcacatagg ccttggagcc gtacataaac
     2821 tgtgggctca ggatgtccca agcgaatggc agaggcccgc cctttgtcac tttcagcttg
     2881 gctgtctggg tgccctcgta gggccgtccc tctccttctc cctcaatttc gaactcgtgg
     2941 ccgttcatgc ttccttccat tctcacctta aaccgcatga actctttgat cacttcctcg
     3001 cccttgctca ccatggtggc ggtagctagc ataacttcgt ataaagtatc ctatacgaag
     3061 ttatttgcct taacccagaa attatcactg ttattcttta gaatggtgca aagaataact
     3121 tcgtataatg tatgctatac gaagttatga attcgatatc aagcttatcg ataatcaacc
     3181 tctggattac aaaatttgtg aaagattgac tggtattctt aactatgttg ctccttttac
     3241 gctatgtgga tacgctgctt taatgccttt gtatcatgct attgcttccc gtatggcttt
     3301 cattttctcc tccttgtata aatcctggtt gctgtctctt tatgaggagt tgtggcccgt
     3361 tgtcaggcaa cgtggcgtgg tgtgcactgt gtttgctgac gcaaccccca ctggttgggg
     3421 cattgccacc acctgtcagc tcctttccgg gactttcgct ttccccctcc ctattgccac
     3481 ggcggaactc atcgccgcct gccttgcccg ctgctggaca ggggctcggc tgttgggcac
     3541 tgacaattcc gtggtgttgt cggggaaatc atcgtccttt ccttggctgc tcgcctatgt
     3601 tgccacctgg attctgcgcg ggacgtcctt ctgctacgtc ccttcggccc tcaatccagc
     3661 ggaccttcct tcccgcggcc tgctgccggc tctgcggcct cttccgcgtc ttcgccttcg
     3721 ccctcagacg agtcggatct ccctttgggc cgcctccccg catcgatacc gagcgctgct
     3781 cgagagatct acgggtggca tccctgtgac ccctccccag tgcctctcct ggccctggaa
     3841 gttgccactc cagtgcccac cagccttgtc ctaataaaat taagttgcat cattttgtct
     3901 gactaggtgt ccttctataa tattatgggg tggagggggg tggtatggag caaggggcaa
     3961 gttgggaaga caacctgtag ggcctgcggg gtctattggg aaccaagctg gagtgcagtg
     4021 gcacaatctt ggctcactgc aatctccgcc tcctgggttc aagcgattct cctgcctcag
     4081 cctcccgagt tgttgggatt ccaggcatgc atgaccaggc tcagctaatt tttgtttttt
     4141 tggtagagac ggggtttcac catattggcc aggctggtct ccaactccta atctcaggtg
     4201 atctacccac cttggcctcc caaattgctg ggattacagg cgtgaaccac tgctcccttc
     4261 cctgtccttc tgattttgta ggtaaccacg tgcggaccga gcggccgcag gaacccctag
     4321 tgatggagtt ggccactccc tctctgcgcg ctcgctcgct cactgaggcc gggcgaccaa
     4381 aggtcgcccg acgcccgggc tttgcccggg cggcctcagt gagcgagcga gcgcgcagct
     4441 gcctgcaggg gcgcctgatg cggtattttc tccttacgca tctgtgcggt atttcacacc
     4501 gcatacgtca aagcaaccat agtacgcgcc ctgtagcggc gcattaagcg cggcgggtgt
     4561 ggtggttacg cgcagcgtga ccgctacact tgccagcgcc ttagcgcccg ctcctttcgc
     4621 tttcttccct tcctttctcg ccacgttcgc cggctttccc cgtcaagctc taaatcgggg
     4681 gctcccttta gggttccgat ttagtgcttt acggcacctc gaccccaaaa aacttgattt
     4741 gggtgatggt tcacgtagtg ggccatcgcc ctgatagacg gtttttcgcc ctttgacgtt
     4801 ggagtccacg ttctttaata gtggactctt gttccaaact ggaacaacac tcaactctat
     4861 ctcgggctat tcttttgatt tataagggat tttgccgatt tcggtctatt ggttaaaaaa
     4921 tgagctgatt taacaaaaat ttaacgcgaa ttttaacaaa atattaacgt ttacaatttt
     4981 atggtgcact ctcagtacaa tctgctctga tgccgcatag ttaagccagc cccgacaccc
     5041 gccaacaccc gctgacgcgc cctgacgggc ttgtctgctc ccggcatccg cttacagaca
     5101 agctgtgacc gtctccggga gctgcatgtg tcagaggttt tcaccgtcat caccgaaacg
     5161 cgcgagacga aagggcctcg tgatacgcct atttttatag gttaatgtca tgataataat
     5221 ggtttcttag acgtcaggtg gcacttttcg gggaaatgtg cgcggaaccc ctatttgttt
     5281 atttttctaa atacattcaa atatgtatcc gctcatgaga caataaccct gataaatgct
     5341 tcaataatat tgaaaaagga agagtatgag tattcaacat ttccgtgtcg cccttattcc
     5401 cttttttgcg gcattttgcc ttcctgtttt tgctcaccca gaaacgctgg tgaaagtaaa
     5461 agatgctgaa gatcagttgg gtgcacgagt gggttacatc gaactggatc tcaacagcgg
     5521 taagatcctt gagagttttc gccccgaaga acgttttcca atgatgagca cttttaaagt
     5581 tctgctatgt ggcgcggtat tatcccgtat tgacgccggg caagagcaac tcggtcgccg
     5641 catacactat tctcagaatg acttggttga gtactcacca gtcacagaaa agcatcttac
     5701 ggatggcatg acagtaagag aattatgcag tgctgccata accatgagtg ataacactgc
     5761 ggccaactta cttctgacaa cgatcggagg accgaaggag ctaaccgctt ttttgcacaa
     5821 catgggggat catgtaactc gccttgatcg ttgggaaccg gagctgaatg aagccatacc
     5881 aaacgacgag cgtgacacca cgatgcctgt agcaatggca acaacgttgc gcaaactatt
     5941 aactggcgaa ctacttactc tagcttcccg gcaacaatta atagactgga tggaggcgga
     6001 taaagttgca ggaccacttc tgcgctcggc ccttccggct ggctggttta ttgctgataa
     6061 atctggagcc ggtgagcgtg ggtctcgcgg tatcattgca gcactggggc cagatggtaa
     6121 gccctcccgt atcgtagtta tctacacgac ggggagtcag gcaactatgg atgaacgaaa
     6181 tagacagatc gctgagatag gtgcctcact gattaagcat tggtaactgt cagaccaagt
     6241 ttactcatat atactttaga ttgatttaaa acttcatttt taatttaaaa ggatctaggt
     6301 gaagatcctt tttgataatc tcatgaccaa aatcccttaa cgtgagtttt cgttccactg
     6361 agcgtcagac cccgtagaaa agatcaaagg atcttcttga gatccttttt ttctgcgcgt
     6421 aatctgctgc ttgcaaacaa aaaaaccacc gctaccagcg gtggtttgtt tgccggatca
     6481 agagctacca actctttttc cgaaggtaac tggcttcagc agagcgcaga taccaaatac
     6541 tgttcttcta gtgtagccgt agttaggcca ccacttcaag aactctgtag caccgcctac
     6601 atacctcgct ctgctaatcc tgttaccagt ggctgctgcc agtggcgata agtcgtgtct
     6661 taccgggttg gactcaagac gatagttacc ggataaggcg cagcggtcgg gctgaacggg
     6721 gggttcgtgc acacagccca gcttggagcg aacgacctac accgaactga gatacctaca
     6781 gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga aaggcggaca ggtatccggt
     6841 aagcggcagg gtcggaacag gagagcgcac gagggagctt ccagggggaa acgcctggta
     6901 tctttatagt cctgtcgggt ttcgccacct ctgacttgag cgtcgatttt tgtgatgctc
     6961 gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg gcctttttac ggttcctggc
     7021 cttttgctgg ccttttgctc acatg
//
