LOCUS       sequence-401143-        7111 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expresses somatically enriched ASAP4b 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 7111)
  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 7111)
  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-ASAP4b-Kv-WPRE
FEATURES             Location/Qualifiers
     source          1..7111
                     /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      3080..3089
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    3096..3129
                     /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    3180..3213
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3238..3826
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3291..3311)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(3709..3720)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    3858..4334
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4057..4076)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4374..4514
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4374..4503
                     /label=AAV2 ITR
     rep_origin      4589..5044
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4676..4695)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4886..4907
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5061..5080)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5180..5202
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5240..5258)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5326..5430
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(5431..5499,5500..6291)
                     /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(5649..5668)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      6462..7050
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6951..6970
                     /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 gctgacctat 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 catggtggcg ctagcataac ttcgtataaa gtatcctata
     3121 cgaagttatt tgccttaacc cagaaattat cactgttatt ctttagaatg gtgcaaagaa
     3181 taacttcgta taatgtatgc tatacgaagt tatgaattcg atatcaagct tatcgataat
     3241 caacctctgg attacaaaat ttgtgaaaga ttgactggta ttcttaacta tgttgctcct
     3301 tttacgctat gtggatacgc tgctttaatg cctttgtatc atgctattgc ttcccgtatg
     3361 gctttcattt tctcctcctt gtataaatcc tggttgctgt ctctttatga ggagttgtgg
     3421 cccgttgtca ggcaacgtgg cgtggtgtgc actgtgtttg ctgacgcaac ccccactggt
     3481 tggggcattg ccaccacctg tcagctcctt tccgggactt tcgctttccc cctccctatt
     3541 gccacggcgg aactcatcgc cgcctgcctt gcccgctgct ggacaggggc tcggctgttg
     3601 ggcactgaca attccgtggt gttgtcgggg aaatcatcgt cctttccttg gctgctcgcc
     3661 tatgttgcca cctggattct gcgcgggacg tccttctgct acgtcccttc ggccctcaat
     3721 ccagcggacc ttccttcccg cggcctgctg ccggctctgc ggcctcttcc gcgtcttcgc
     3781 cttcgccctc agacgagtcg gatctccctt tgggccgcct ccccgcatcg ataccgagcg
     3841 ctgctcgaga gatctacggg tggcatccct gtgacccctc cccagtgcct ctcctggccc
     3901 tggaagttgc cactccagtg cccaccagcc ttgtcctaat aaaattaagt tgcatcattt
     3961 tgtctgacta ggtgtccttc tataatatta tggggtggag gggggtggta tggagcaagg
     4021 ggcaagttgg gaagacaacc tgtagggcct gcggggtcta ttgggaacca agctggagtg
     4081 cagtggcaca atcttggctc actgcaatct ccgcctcctg ggttcaagcg attctcctgc
     4141 ctcagcctcc cgagttgttg ggattccagg catgcatgac caggctcagc taatttttgt
     4201 ttttttggta gagacggggt ttcaccatat tggccaggct ggtctccaac tcctaatctc
     4261 aggtgatcta cccaccttgg cctcccaaat tgctgggatt acaggcgtga accactgctc
     4321 ccttccctgt ccttctgatt ttgtaggtaa ccacgtgcgg accgagcggc cgcaggaacc
     4381 cctagtgatg gagttggcca ctccctctct gcgcgctcgc tcgctcactg aggccgggcg
     4441 accaaaggtc gcccgacgcc cgggctttgc ccgggcggcc tcagtgagcg agcgagcgcg
     4501 cagctgcctg caggggcgcc tgatgcggta ttttctcctt acgcatctgt gcggtatttc
     4561 acaccgcata cgtcaaagca accatagtac gcgccctgta gcggcgcatt aagcgcggcg
     4621 ggtgtggtgg ttacgcgcag cgtgaccgct acacttgcca gcgccttagc gcccgctcct
     4681 ttcgctttct tcccttcctt tctcgccacg ttcgccggct ttccccgtca agctctaaat
     4741 cgggggctcc ctttagggtt ccgatttagt gctttacggc acctcgaccc caaaaaactt
     4801 gatttgggtg atggttcacg tagtgggcca tcgccctgat agacggtttt tcgccctttg
     4861 acgttggagt ccacgttctt taatagtgga ctcttgttcc aaactggaac aacactcaac
     4921 tctatctcgg gctattcttt tgatttataa gggattttgc cgatttcggt ctattggtta
     4981 aaaaatgagc tgatttaaca aaaatttaac gcgaatttta acaaaatatt aacgtttaca
     5041 attttatggt gcactctcag tacaatctgc tctgatgccg catagttaag ccagccccga
     5101 cacccgccaa cacccgctga cgcgccctga cgggcttgtc tgctcccggc atccgcttac
     5161 agacaagctg tgaccgtctc cgggagctgc atgtgtcaga ggttttcacc gtcatcaccg
     5221 aaacgcgcga gacgaaaggg cctcgtgata cgcctatttt tataggttaa tgtcatgata
     5281 ataatggttt cttagacgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt
     5341 tgtttatttt tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa
     5401 atgcttcaat aatattgaaa aaggaagagt atgagtattc aacatttccg tgtcgccctt
     5461 attccctttt ttgcggcatt ttgccttcct gtttttgctc acccagaaac gctggtgaaa
     5521 gtaaaagatg ctgaagatca gttgggtgca cgagtgggtt acatcgaact ggatctcaac
     5581 agcggtaaga tccttgagag ttttcgcccc gaagaacgtt ttccaatgat gagcactttt
     5641 aaagttctgc tatgtggcgc ggtattatcc cgtattgacg ccgggcaaga gcaactcggt
     5701 cgccgcatac actattctca gaatgacttg gttgagtact caccagtcac agaaaagcat
     5761 cttacggatg gcatgacagt aagagaatta tgcagtgctg ccataaccat gagtgataac
     5821 actgcggcca acttacttct gacaacgatc ggaggaccga aggagctaac cgcttttttg
     5881 cacaacatgg gggatcatgt aactcgcctt gatcgttggg aaccggagct gaatgaagcc
     5941 ataccaaacg acgagcgtga caccacgatg cctgtagcaa tggcaacaac gttgcgcaaa
     6001 ctattaactg gcgaactact tactctagct tcccggcaac aattaataga ctggatggag
     6061 gcggataaag ttgcaggacc acttctgcgc tcggcccttc cggctggctg gtttattgct
     6121 gataaatctg gagccggtga gcgtgggtct cgcggtatca ttgcagcact ggggccagat
     6181 ggtaagccct cccgtatcgt agttatctac acgacgggga gtcaggcaac tatggatgaa
     6241 cgaaatagac agatcgctga gataggtgcc tcactgatta agcattggta actgtcagac
     6301 caagtttact catatatact ttagattgat ttaaaacttc atttttaatt taaaaggatc
     6361 taggtgaaga tcctttttga taatctcatg accaaaatcc cttaacgtga gttttcgttc
     6421 cactgagcgt cagaccccgt agaaaagatc aaaggatctt cttgagatcc tttttttctg
     6481 cgcgtaatct gctgcttgca aacaaaaaaa ccaccgctac cagcggtggt ttgtttgccg
     6541 gatcaagagc taccaactct ttttccgaag gtaactggct tcagcagagc gcagatacca
     6601 aatactgttc ttctagtgta gccgtagtta ggccaccact tcaagaactc tgtagcaccg
     6661 cctacatacc tcgctctgct aatcctgtta ccagtggctg ctgccagtgg cgataagtcg
     6721 tgtcttaccg ggttggactc aagacgatag ttaccggata aggcgcagcg gtcgggctga
     6781 acggggggtt cgtgcacaca gcccagcttg gagcgaacga cctacaccga actgagatac
     6841 ctacagcgtg agctatgaga aagcgccacg cttcccgaag ggagaaaggc ggacaggtat
     6901 ccggtaagcg gcagggtcgg aacaggagag cgcacgaggg agcttccagg gggaaacgcc
     6961 tggtatcttt atagtcctgt cgggtttcgc cacctctgac ttgagcgtcg atttttgtga
     7021 tgctcgtcag gggggcggag cctatggaaa aacgccagca acgcggcctt tttacggttc
     7081 ctggcctttt gctggccttt tgctcacatg t
//
