LOCUS       sequence-477842-        7033 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cre dependent Green fluorescent, negative response-polarity voltage 
            indicator for mammalian expression.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7033)
  AUTHORS   Haziza S, Chrapkiewicz R, Zhang Y, Kruzhilin V, Li J, Li J, Delamare
            G, Swanson R, Buzsaki G, Kannan M, Vasan G, Lin MZ, Zeng H, Daigle 
            TL, Schnitzer MJ
  TITLE     Imaging high-frequency voltage dynamics in multiple neuron classes 
            of behaving mammals.
  JOURNAL   Cell, 188, 1-23, August 7, 2025. doi: 10.1016/j.cell.2025.06.028
  PUBMED    39185175
REFERENCE   2  (bases 1 to 7033)
  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: "Cell, 188, 
            1-23, August 7, 2025. doi: 10.1016/j.cell.2025.06.028"
COMMENT     Sequence Label: pAAV-EF1a-DIO-Ace-mNeon2-WPRE
FEATURES             Location/Qualifiers
     source          1..7033
                     /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      2093..2102
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     regulatory      3002..3011
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3018..3051
                     /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    3102..3135
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3160..3748
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3213..3233)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3631..3642)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3780..4256
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(3979..3998)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4296..4436
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4296..4425
                     /label=AAV2 ITR
     rep_origin      4511..4966
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4598..4617)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4808..4829
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(4983..5002)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5102..5124
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5162..5180)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5248..5352
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5353..5421,5422..6213)
                     /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(5571..5590)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6384..6972
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6873..6892
                     /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 gcgccttaca cctcgttctc gtagcagaac acgttgatgt cgatctggtc cagggggatg
     1621 tactcgccct cgctggtgat cctgctcttc ttgtacagct cgtccatgcc catcacatcg
     1681 gtaaaggcct tttgccactc cttgaagttg agctcggtct tggagtgctt gagctccgtc
     1741 ttacggaaca cgtacatcgg ctggttcttc agatagttag ccgccattgg cttggcaaag
     1801 gtgtaggtgg tccgcgcagt gctcctgtag cgcttgccat ttccagtggt gtaactccac
     1861 ttaaaggtac tgatgatggt tttgtcgttg gggtaagtct tcttcgacct gcaccagtcc
     1921 gcagcggtca gcgagttggt catcacagga ccgtcagcag ggaaaccagt ccccttcacc
     1981 tgggcctctc ctttgatgtg gcttccctcg taggtgtagc ggtagttaac agtaagggag
     2041 gcaccatctt caaactgcat tgtgcgatgg acttggtatc cggagccatc taccatggcg
     2101 gcctggaaag gcgacatccc gtcagggtag ggcaggtact gatggaagcc atacccgata
     2161 tgagggacca gaatccaggg ggagaactgg aggtcaccct tggtggactt caggtttaac
     2221 tcctcataac catcatttgg attgccggtg ccctgaccca ccatgtcaaa gtccacaccg
     2281 ttgatggagc caaagatgtg taactcatgt gtcgctgggt aggacttgaa gatagtctca
     2341 tgggcaatga ggcaccccat tccaaacagt ggcttagcga gcacatccag gattcccatg
     2401 agaatggcct cattgtcgac ggacagcacg ccactaccag acccgaaagc ccacacgaca
     2461 ggataaaaca cccagaggat tgacaggaag acgagcagac tgacatagga gctatggtac
     2521 tcggggcttt tcttcagatc gtctttccaa gtggcgttat acatcccgta tgtcaggact
     2581 gcctgcatca cacatccagc gataaagtac acccatttga acttgtgttc atcctcgaag
     2641 gcgcccagaa taccaaacac gatcatgaaa atgtctgcgc cgatgaccca agaaatcatc
     2701 actccgccca tcttggtcat gacgatgaga tcgagcagga gcagtggtgt ggtcagcacc
     2761 cagctaatat atcttgcgta gaccacctgc ctttcaccat tcattgtcag gtcctggtag
     2821 ttcacggcca ttgcatagta agcggtagcg gcgatagtgc agatggcaat gttgatatag
     2881 tatgcgctct tcagctctga aggccgaaag tgcacagcca ctccaaaagc aatagcagca
     2941 gcagccataa tagcaaagac aatccactgt gcaatcatgc cggtctcggt ttccacgtca
     3001 gccatggtgg cgctagcata acttcgtata aagtatccta tacgaagtta tttgccttaa
     3061 cccagaaatt atcactgtta ttctttagaa tggtgcaaag aataacttcg tataatgtat
     3121 gctatacgaa gttatgaatt cgatatcaag cttatcgata atcaacctct ggattacaaa
     3181 atttgtgaaa gattgactgg tattcttaac tatgttgctc cttttacgct atgtggatac
     3241 gctgctttaa tgcctttgta tcatgctatt gcttcccgta tggctttcat tttctcctcc
     3301 ttgtataaat cctggttgct gtctctttat gaggagttgt ggcccgttgt caggcaacgt
     3361 ggcgtggtgt gcactgtgtt tgctgacgca acccccactg gttggggcat tgccaccacc
     3421 tgtcagctcc tttccgggac tttcgctttc cccctcccta ttgccacggc ggaactcatc
     3481 gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt tgggcactga caattccgtg
     3541 gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg cctatgttgc cacctggatt
     3601 ctgcgcggga cgtccttctg ctacgtccct tcggccctca atccagcgga ccttccttcc
     3661 cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc gccttcgccc tcagacgagt
     3721 cggatctccc tttgggccgc ctccccgcat cgataccgag cgctgctcga gagatctacg
     3781 ggtggcatcc ctgtgacccc tccccagtgc ctctcctggc cctggaagtt gccactccag
     3841 tgcccaccag ccttgtccta ataaaattaa gttgcatcat tttgtctgac taggtgtcct
     3901 tctataatat tatggggtgg aggggggtgg tatggagcaa ggggcaagtt gggaagacaa
     3961 cctgtagggc ctgcggggtc tattgggaac caagctggag tgcagtggca caatcttggc
     4021 tcactgcaat ctccgcctcc tgggttcaag cgattctcct gcctcagcct cccgagttgt
     4081 tgggattcca ggcatgcatg accaggctca gctaattttt gtttttttgg tagagacggg
     4141 gtttcaccat attggccagg ctggtctcca actcctaatc tcaggtgatc tacccacctt
     4201 ggcctcccaa attgctggga ttacaggcgt gaaccactgc tcccttccct gtccttctga
     4261 ttttgtaggt aaccacgtgc ggaccgagcg gccgcaggaa cccctagtga tggagttggc
     4321 cactccctct ctgcgcgctc gctcgctcac tgaggccggg cgaccaaagg tcgcccgacg
     4381 cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg cgcagctgcc tgcaggggcg
     4441 cctgatgcgg tattttctcc ttacgcatct gtgcggtatt tcacaccgca tacgtcaaag
     4501 caaccatagt acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc
     4561 agcgtgaccg ctacacttgc cagcgcctta gcgcccgctc ctttcgcttt cttcccttcc
     4621 tttctcgcca cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg
     4681 ttccgattta gtgctttacg gcacctcgac cccaaaaaac ttgatttggg tgatggttca
     4741 cgtagtgggc catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc
     4801 tttaatagtg gactcttgtt ccaaactgga acaacactca actctatctc gggctattct
     4861 tttgatttat aagggatttt gccgatttcg gtctattggt taaaaaatga gctgatttaa
     4921 caaaaattta acgcgaattt taacaaaata ttaacgttta caattttatg gtgcactctc
     4981 agtacaatct gctctgatgc cgcatagtta agccagcccc gacacccgcc aacacccgct
     5041 gacgcgccct gacgggcttg tctgctcccg gcatccgctt acagacaagc tgtgaccgtc
     5101 tccgggagct gcatgtgtca gaggttttca ccgtcatcac cgaaacgcgc gagacgaaag
     5161 ggcctcgtga tacgcctatt tttataggtt aatgtcatga taataatggt ttcttagacg
     5221 tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta tttgtttatt tttctaaata
     5281 cattcaaata tgtatccgct catgagacaa taaccctgat aaatgcttca ataatattga
     5341 aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc ttattccctt ttttgcggca
     5401 ttttgccttc ctgtttttgc tcacccagaa acgctggtga aagtaaaaga tgctgaagat
     5461 cagttgggtg cacgagtggg ttacatcgaa ctggatctca acagcggtaa gatccttgag
     5521 agttttcgcc ccgaagaacg ttttccaatg atgagcactt ttaaagttct gctatgtggc
     5581 gcggtattat cccgtattga cgccgggcaa gagcaactcg gtcgccgcat acactattct
     5641 cagaatgact tggttgagta ctcaccagtc acagaaaagc atcttacgga tggcatgaca
     5701 gtaagagaat tatgcagtgc tgccataacc atgagtgata acactgcggc caacttactt
     5761 ctgacaacga tcggaggacc gaaggagcta accgcttttt tgcacaacat gggggatcat
     5821 gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag ccataccaaa cgacgagcgt
     5881 gacaccacga tgcctgtagc aatggcaaca acgttgcgca aactattaac tggcgaacta
     5941 cttactctag cttcccggca acaattaata gactggatgg aggcggataa agttgcagga
     6001 ccacttctgc gctcggccct tccggctggc tggtttattg ctgataaatc tggagccggt
     6061 gagcgtgggt ctcgcggtat cattgcagca ctggggccag atggtaagcc ctcccgtatc
     6121 gtagttatct acacgacggg gagtcaggca actatggatg aacgaaatag acagatcgct
     6181 gagataggtg cctcactgat taagcattgg taactgtcag accaagttta ctcatatata
     6241 ctttagattg atttaaaact tcatttttaa tttaaaagga tctaggtgaa gatccttttt
     6301 gataatctca tgaccaaaat cccttaacgt gagttttcgt tccactgagc gtcagacccc
     6361 gtagaaaaga tcaaaggatc ttcttgagat cctttttttc tgcgcgtaat ctgctgcttg
     6421 caaacaaaaa aaccaccgct accagcggtg gtttgtttgc cggatcaaga gctaccaact
     6481 ctttttccga aggtaactgg cttcagcaga gcgcagatac caaatactgt tcttctagtg
     6541 tagccgtagt taggccacca cttcaagaac tctgtagcac cgcctacata cctcgctctg
     6601 ctaatcctgt taccagtggc tgctgccagt ggcgataagt cgtgtcttac cgggttggac
     6661 tcaagacgat agttaccgga taaggcgcag cggtcgggct gaacgggggg ttcgtgcaca
     6721 cagcccagct tggagcgaac gacctacacc gaactgagat acctacagcg tgagctatga
     6781 gaaagcgcca cgcttcccga agggagaaag gcggacaggt atccggtaag cggcagggtc
     6841 ggaacaggag agcgcacgag ggagcttcca gggggaaacg cctggtatct ttatagtcct
     6901 gtcgggtttc gccacctctg acttgagcgt cgatttttgt gatgctcgtc aggggggcgg
     6961 agcctatgga aaaacgccag caacgcggcc tttttacggt tcctggcctt ttgctggcct
     7021 tttgctcaca tgt
//
