LOCUS       sequence-401146-        7112 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses somatically enriched ASAP4e from the EF-1alpha promoter in
            a cre-dependent manner.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7112)
  AUTHORS   Evans SW, Shi DQ, Chavarha M, Plitt MH, Taxidis J, Madruga B, Fan 
            JL, Hwang FJ, van Keulen SC, Suomivuori CM, Pang MM, Su S, Lee S, 
            Hao YA, Zhang G, Jiang D, Pradhan L, Roth RH, Liu Y, Dorian CC, 
            Reese AL, Negrean A, Losonczy A, Makinson CD, Wang S, Clandinin TR, 
            Dror RO, Ding JB, Ji N, Golshani P, Giocomo LM, Bi GQ, Lin MZ
  TITLE     A positively tuned voltage indicator for extended electrical 
            recordings in the brain.
  JOURNAL   Nat Methods. 2023 Jul;20(7):1104-1113. doi: 
            10.1038/s41592-023-01913-z. Epub 2023 Jul 6.
  PUBMED    37429962
REFERENCE   2  (bases 1 to 7112)
  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/s41592-023-01913-z"; journalName: "Nat Methods"; date: 
            "2023-07"; volume: "20"; issue: "7"; pages: "1104-1113"
COMMENT     Sequence Label: AAV-EF1a-FLEx-ASAP4e-Kv-WPRE
FEATURES             Location/Qualifiers
     source          1..7112
                     /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)."
     protein_bind    3097..3130
                     /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    3181..3214
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3239..3827
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3292..3312)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3710..3721)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3859..4335
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4058..4077)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4375..4515
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4375..4504
                     /label=AAV2 ITR
     rep_origin      4590..5045
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4677..4696)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4887..4908
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5062..5081)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5181..5203
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5241..5259)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5327..5431
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5432..5500,5501..6292)
                     /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(5650..5669)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6463..7051
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6952..6971
                     /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 gcgcctcatt aaaagcgggt cgcttccgga aaatcggtcg cgcagctaat aaagctatca
     1621 atgctgctca tgctgcgcat atcaatcacg ccttcggtgc gcgccggcag cggcgccacc
     1681 gggctcggca tgctgctcat ttccagttct tccggcggtt tgctctgcgg cgccatttct
     1741 ttggtgttca gaatcggctg gctctgacgc gtggaagatc cagatgaacc tgaagaaccc
     1801 ttaagggtta ccacttcaag ttgtttcttc tgtgaagcca gctgcattat tcttattaag
     1861 ataacaattc tcagaactcg aagaagagta accatctgcg gcatctgatc tgtgttatat
     1921 tccagtttat gccccagaat gttgccatct tctttaaaat cggtgccttt cagttcaatg
     1981 cggttcacca gggtatcgcc ttcaaatttc accaccgcgc gggttttata tttgccatca
     2041 tctttaaagc taatggtgcg ttcctgcaca tagccttccg gcatcgcgct tttaaaaaaa
     2101 tcatggcgtt tcatatgatc cggatagcgg ctaaagcact gcacgccata ggttaaggtg
     2161 gtcaccaggg tcggccacgg caccggtagt ttgccggtgg tgcaaataaa tttcagggtc
     2221 agtttgccaa tggtcgcatc gccttcgcct tcgccgcgca cgctaaattt atggccgttc
     2281 acatcgccat ccagttccac cagaatcggc accacgccgg taaacagttc ttcgccctgg
     2341 ctcgcgctgc cgccggtgcc gccatacagt tcatccatgc cgtgtgtaat gcccgctgcg
     2401 gtcacaaatt ccagcagcac catgtgatcg cgtttttcgt tcggatcttt gctcagcacg
     2461 gtctggtggc tcagataatg gttatccggc agcagcaccg ggccatcgcc aatcggggtg
     2521 ttctgctgat aatgatccgc cagctgcacg ctgccatctt ccacgttatg gcgcacggta
     2581 aagttcgctt taatgccgtt tttctgttta tccgcggtaa tatacacgtt accatcaacg
     2641 ggtgtaccgt cagagaagga gtaggtcata ttaattagca gagtgcccac tactatgact
     2701 gcatccaaag tattcagttt ggaccggaaa tagttcttga agccttcaac aaacactctc
     2761 atgagaacat caacaaggaa gaagagtgct atagccaggg aaacaccttc aagaatctct
     2821 ctaatgcctc ttttcttctc actgatggcc agatccacaa tcaccactat gatgtctaca
     2881 atgataagca caactccaaa tactctgaat ccaaatgaca tcacaaacgg ggaaattttt
     2941 ctcctaatgg tattggaaca gctgtcttgt tcattaccct catcacgacc atcatcaatc
     3001 tttatcgtgg gctcatctgc tgttggagag ctcgaagttt tagtgagctc tgacccctgt
     3061 tcatacctca cagtcgtctc catggtgggc gctagcataa cttcgtataa agtatcctat
     3121 acgaagttat ttgccttaac ccagaaatta tcactgttat tctttagaat ggtgcaaaga
     3181 ataacttcgt ataatgtatg ctatacgaag ttatgaattc gatatcaagc ttatcgataa
     3241 tcaacctctg gattacaaaa tttgtgaaag attgactggt attcttaact atgttgctcc
     3301 ttttacgcta tgtggatacg ctgctttaat gcctttgtat catgctattg cttcccgtat
     3361 ggctttcatt ttctcctcct tgtataaatc ctggttgctg tctctttatg aggagttgtg
     3421 gcccgttgtc aggcaacgtg gcgtggtgtg cactgtgttt gctgacgcaa cccccactgg
     3481 ttggggcatt gccaccacct gtcagctcct ttccgggact ttcgctttcc ccctccctat
     3541 tgccacggcg gaactcatcg ccgcctgcct tgcccgctgc tggacagggg ctcggctgtt
     3601 gggcactgac aattccgtgg tgttgtcggg gaaatcatcg tcctttcctt ggctgctcgc
     3661 ctatgttgcc acctggattc tgcgcgggac gtccttctgc tacgtccctt cggccctcaa
     3721 tccagcggac cttccttccc gcggcctgct gccggctctg cggcctcttc cgcgtcttcg
     3781 ccttcgccct cagacgagtc ggatctccct ttgggccgcc tccccgcatc gataccgagc
     3841 gctgctcgag agatctacgg gtggcatccc tgtgacccct ccccagtgcc tctcctggcc
     3901 ctggaagttg ccactccagt gcccaccagc cttgtcctaa taaaattaag ttgcatcatt
     3961 ttgtctgact aggtgtcctt ctataatatt atggggtgga ggggggtggt atggagcaag
     4021 gggcaagttg ggaagacaac ctgtagggcc tgcggggtct attgggaacc aagctggagt
     4081 gcagtggcac aatcttggct cactgcaatc tccgcctcct gggttcaagc gattctcctg
     4141 cctcagcctc ccgagttgtt gggattccag gcatgcatga ccaggctcag ctaatttttg
     4201 tttttttggt agagacgggg tttcaccata ttggccaggc tggtctccaa ctcctaatct
     4261 caggtgatct acccaccttg gcctcccaaa ttgctgggat tacaggcgtg aaccactgct
     4321 cccttccctg tccttctgat tttgtaggta accacgtgcg gaccgagcgg ccgcaggaac
     4381 ccctagtgat ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc
     4441 gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc
     4501 gcagctgcct gcaggggcgc ctgatgcggt attttctcct tacgcatctg tgcggtattt
     4561 cacaccgcat acgtcaaagc aaccatagta cgcgccctgt agcggcgcat taagcgcggc
     4621 gggtgtggtg gttacgcgca gcgtgaccgc tacacttgcc agcgccttag cgcccgctcc
     4681 tttcgctttc ttcccttcct ttctcgccac gttcgccggc tttccccgtc aagctctaaa
     4741 tcgggggctc cctttagggt tccgatttag tgctttacgg cacctcgacc ccaaaaaact
     4801 tgatttgggt gatggttcac gtagtgggcc atcgccctga tagacggttt ttcgcccttt
     4861 gacgttggag tccacgttct ttaatagtgg actcttgttc caaactggaa caacactcaa
     4921 ctctatctcg ggctattctt ttgatttata agggattttg ccgatttcgg tctattggtt
     4981 aaaaaatgag ctgatttaac aaaaatttaa cgcgaatttt aacaaaatat taacgtttac
     5041 aattttatgg tgcactctca gtacaatctg ctctgatgcc gcatagttaa gccagccccg
     5101 acacccgcca acacccgctg acgcgccctg acgggcttgt ctgctcccgg catccgctta
     5161 cagacaagct gtgaccgtct ccgggagctg catgtgtcag aggttttcac cgtcatcacc
     5221 gaaacgcgcg agacgaaagg gcctcgtgat acgcctattt ttataggtta atgtcatgat
     5281 aataatggtt tcttagacgt caggtggcac ttttcgggga aatgtgcgcg gaacccctat
     5341 ttgtttattt ttctaaatac attcaaatat gtatccgctc atgagacaat aaccctgata
     5401 aatgcttcaa taatattgaa aaaggaagag tatgagtatt caacatttcc gtgtcgccct
     5461 tattcccttt tttgcggcat tttgccttcc tgtttttgct cacccagaaa cgctggtgaa
     5521 agtaaaagat gctgaagatc agttgggtgc acgagtgggt tacatcgaac tggatctcaa
     5581 cagcggtaag atccttgaga gttttcgccc cgaagaacgt tttccaatga tgagcacttt
     5641 taaagttctg ctatgtggcg cggtattatc ccgtattgac gccgggcaag agcaactcgg
     5701 tcgccgcata cactattctc agaatgactt ggttgagtac tcaccagtca cagaaaagca
     5761 tcttacggat ggcatgacag taagagaatt atgcagtgct gccataacca tgagtgataa
     5821 cactgcggcc aacttacttc tgacaacgat cggaggaccg aaggagctaa ccgctttttt
     5881 gcacaacatg ggggatcatg taactcgcct tgatcgttgg gaaccggagc tgaatgaagc
     5941 cataccaaac gacgagcgtg acaccacgat gcctgtagca atggcaacaa cgttgcgcaa
     6001 actattaact ggcgaactac ttactctagc ttcccggcaa caattaatag actggatgga
     6061 ggcggataaa gttgcaggac cacttctgcg ctcggccctt ccggctggct ggtttattgc
     6121 tgataaatct ggagccggtg agcgtgggtc tcgcggtatc attgcagcac tggggccaga
     6181 tggtaagccc tcccgtatcg tagttatcta cacgacgggg agtcaggcaa ctatggatga
     6241 acgaaataga cagatcgctg agataggtgc ctcactgatt aagcattggt aactgtcaga
     6301 ccaagtttac tcatatatac tttagattga tttaaaactt catttttaat ttaaaaggat
     6361 ctaggtgaag atcctttttg ataatctcat gaccaaaatc ccttaacgtg agttttcgtt
     6421 ccactgagcg tcagaccccg tagaaaagat caaaggatct tcttgagatc ctttttttct
     6481 gcgcgtaatc tgctgcttgc aaacaaaaaa accaccgcta ccagcggtgg tttgtttgcc
     6541 ggatcaagag ctaccaactc tttttccgaa ggtaactggc ttcagcagag cgcagatacc
     6601 aaatactgtt cttctagtgt agccgtagtt aggccaccac ttcaagaact ctgtagcacc
     6661 gcctacatac ctcgctctgc taatcctgtt accagtggct gctgccagtg gcgataagtc
     6721 gtgtcttacc gggttggact caagacgata gttaccggat aaggcgcagc ggtcgggctg
     6781 aacggggggt tcgtgcacac agcccagctt ggagcgaacg acctacaccg aactgagata
     6841 cctacagcgt gagctatgag aaagcgccac gcttcccgaa gggagaaagg cggacaggta
     6901 tccggtaagc ggcagggtcg gaacaggaga gcgcacgagg gagcttccag ggggaaacgc
     6961 ctggtatctt tatagtcctg tcgggtttcg ccacctctga cttgagcgtc gatttttgtg
     7021 atgctcgtca ggggggcgga gcctatggaa aaacgccagc aacgcggcct ttttacggtt
     7081 cctggccttt tgctggcctt ttgctcacat gt
//
