LOCUS       sequence-414423-        7713 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Expressing soma-targeted CheRiff_CFP in a Flp-dependent manner.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7713)
  TITLE     AAV constructs for expressing genetically encoded voltage indicators
            in mice
  JOURNAL   Unpublished
REFERENCE   2  (bases 1 to 7713)
  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: 
            "Unpublished"
COMMENT     Sequence Label: HT147_pAAV_EF1a_fDiO_somCheRiff_CFP
FEATURES             Location/Qualifiers
     source          1..7713
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   1..141
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   12..141
                     /label=AAV2 ITR
     promoter        242..1419
                     /label=EF-1-alpha promoter
                     /note="strong constitutive promoter for human elongation 
                     factor EF-1-alpha"
     intron          473..1410
                     /label=EF-1-alpha intron A
                     /note="intron upstream of the start codon of human 
                     EF-1-alpha"
     primer_bind     1367..1387
                     /label=EF1a-F
                     /note="Human elongation factor-1a promoter, forward primer"
     protein_bind    1463..1510
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     CDS             complement(join(1855..2565,2566..2568))
                     /codon_start=1
                     /product="cyan variant of GFP"
                     /label=CFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHRFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTWGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYISHNVYITADKQKNGIKA
                     HFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1897..1918)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     2246..2265
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     2505..2526
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     CDS             complement(2647..3594)
                     /codon_start=1
                     /product="blue light-activated optogenetic actuator derived
                     from Scherffelia dubia channelrhodopsin (Hochbaum et al., 
                     2014)"
                     /label=CheRiff
                     /translation="MGGAPAPDAHSAPPGNDSAGGSEYHAPAGYQVNPPYHPVHGYEEQ
                     CSSIYIYYGALWEQETARGFQWFAVFLSALFLAFYGWHAYKASVGWEEVYVCSVELIKV
                     ILEIYFEFTSPAMLFLYGGNITPWLRYAEWLLTCPVILIHLSNITGLSEAYNKRTMALL
                     VSDLGTICMGVTAALATGWVKWLFYCIGLVYGTQTFYNAGIIYVESYYIMPAGGCKKLV
                     LAMTAVYYSSWLMFPGLFIFGPEGMHTLSVAGSTIGHTIADLLSKNIWGLLGHFLRIKI
                     HEHIIMYGDIRRPVSSQFLGRKVDVLAFVTEEDKV"
     regulatory      3591..3600
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     protein_bind    complement(3639..3686)
                     /label=FRT
                     /bound_moiety="FLP recombinase from the Saccharomyces 
                     cerevisiae 2u plasmid"
                     /note="FLP-mediated recombination occurs in the 8-bp core 
                     sequence TCTAGAAA (Turan and Bode, 2011)."
     misc_feature    3840..4428
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3893..3913)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4311..4322)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     polyA_signal    4460..4936
                     /label=hGH poly(A) signal
                     /note="human growth hormone polyadenylation signal"
     primer_bind     complement(4659..4678)
                     /label=hGH-PA-R
                     /note="Human growth hormone terminator, reverse primer"
     repeat_region   4976..5116
                     /label=AAV2 ITR
                     /note="inverted terminal repeat of adeno-associated virus 
                     serotype 2"
     repeat_region   4976..5105
                     /label=AAV2 ITR
     rep_origin      5191..5646
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5278..5297)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5488..5509
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(5663..5682)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     primer_bind     5782..5804
                     /label=pGEX 3'
                     /note="pGEX vectors, reverse primer"
     primer_bind     complement(5842..5860)
                     /label=pBRforEco
                     /note="pBR322 vectors, upsteam of EcoRI site, forward 
                     primer"
     promoter        5928..6032
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6033..6101,6102..6893)
                     /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(6251..6270)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7064..7652
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7553..7572
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 cctgcaggca gctgcgcgct cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc
       61 gggcgacctt tggtcgcccg gcctcagtga gcgagcgagc gcgcagagag ggagtggcca
      121 actccatcac taggggttcc tgcggccgca cgcgtaagct ttgcaaagat ggataaagtt
      181 ttaaacagag aggaatcttt gcagctaatg gaccttctag gtcttgaaag gagtgggaat
      241 tggctccggt gcccgtcagt gggcagagcg cacatcgccc acagtccccg agaagttggg
      301 gggaggggtc ggcaattgaa ccggtgccta gagaaggtgg cgcggggtaa actgggaaag
      361 tgatgtcgtg tactggctcc gcctttttcc cgagggtggg ggagaaccgt atataagtgc
      421 agtagtcgcc gtgaacgttc tttttcgcaa cgggtttgcc gccagaacac aggtaagtgc
      481 cgtgtgtggt tcccgcgggc ctggcctctt tacgggttat ggcccttgcg tgccttgaat
      541 tacttccact ggctgcagta cgtgattctt gatcccgagc ttcgggttgg aagtgggtgg
      601 gagagttcga ggccttgcgc ttaaggagcc ccttcgcctc gtgcttgagt tgaggcctgg
      661 cctgggcgct ggggccgccg cgtgcgaatc tggtggcacc ttcgcgcctg tctcgctgct
      721 ttcgataagt ctctagccat ttaaaatttt tgatgacctg ctgcgacgct ttttttctgg
      781 caagatagtc ttgtaaatgc gggccaagat ctgcacactg gtatttcggt ttttggggcc
      841 gcgggcggcg acggggcccg tgcgtcccag cgcacatgtt cggcgaggcg gggcctgcga
      901 gcgcggccac cgagaatcgg acgggggtag tctcaagctg gccggcctgc tctggtgcct
      961 ggcctcgcgc cgccgtgtat cgccccgccc tgggcggcaa ggctggcccg gtcggcacca
     1021 gttgcgtgag cggaaagatg gccgcttccc ggccctgctg cagggagctc aaaatggagg
     1081 acgcggcgct cgggagagcg ggcgggtgag tcacccacac aaaggaaaag ggcctttccg
     1141 tcctcagccg tcgcttcatg tgactccacg gagtaccggg cgccgtccag gcacctcgat
     1201 tagttctcga gcttttggag tacgtcgtct ttaggttggg gggaggggtt ttatgcgatg
     1261 gagtttcccc acactgagtg ggtggagact gaagttaggc cagcttggca cttgatgtaa
     1321 ttctccttgg aatttgccct ttttgagttt ggatcttggt tcattctcaa gcctcagaca
     1381 gtggttcaaa gtttttttct tccatttcag gtgtcgtgag gtaccggatc ctctagagtc
     1441 gacggctgtg ggcccgtaac aagaagttcc tattccgaag ttcctattct ctagaaagta
     1501 taggaacttc atcaaaatag gaagaccaat gcttcaccat cgacccgaat tgccaagcat
     1561 caccatcgac agaagttcct attccgaagt tcctattctt caaaaggtat aggaacttcg
     1621 cctcaaagcc gcgtgacgag ctagttacgg cgcgccttag aatctggtgg cttcagggaa
     1681 gtccgtggca cagctgatga agctgtcaat gctggacatg ctccgcatgt cgatgacgcc
     1741 ctccgtgcgt gcgggcagag gggccacggg gctgggcatg ctgctcatct ccagctcctc
     1801 tggaggcttg ctctgcgggg ccatctcctt ggtgttgagg atgggctggg actgcttgta
     1861 cagctcgtcc atgccgagag tgatcccggc ggcggtcacg aactccagca ggaccatgtg
     1921 atcgcgcttc tcgttggggt ctttgctcag ggcggactgg gtgctcaggt agtggttgtc
     1981 gggcagcagc acggggccgt cgccgatggg ggtgttctgc tggtagtggt cggcgagctg
     2041 cacgctgccg tcctcgatgt tgtggcggat cttgaagtgg gccttgatgc cgttcttctg
     2101 cttgtcggcg gtgatataga cgttgtggct gatgtagttg tactccagct tgtgccccag
     2161 gatgttgccg tcctccttga agtcgatgcc cttcagctcg atgcggttca ccagggtgtc
     2221 gccctcgaac ttcacctcgg cgcgggtctt gtagttgccg tcgtccttga agaagatggt
     2281 acgctcctgg acgtagcctt cgggcatggc ggacttgaag aagtcgtgct gcttcatgtg
     2341 gtcggggtag cggctgaagc actgcacgcc ccaggtcagg gtggtcacga gggtgggcca
     2401 gggcacgggc agcttgccgg tggtgcagat gaacttcagg gtcagcttgc cgtaggtggc
     2461 atcgccctcg ccctcgccgg acacgctgaa cctgtggccg tttacgtcgc cgtccagctc
     2521 gaccaggatg ggcaccaccc cggtgaacag ctcctcgccc ttgctcacca ccacgttgat
     2581 gtcgatctgg tccaggggga tgtactcgcc ctcgctggtg atcctgctct tgctcttggc
     2641 ggccgccact ttatcttcct ctgtcacgaa ggccagcacg tccaccttgc gtcccagaaa
     2701 ctgggagctc actggtctcc tgatatcgcc gtacataatg atatgctcgt gaattttgat
     2761 ccgcaggaag tgccccagca gtccccaaat attcttggac agcaggtcgg cgatggtatg
     2821 gccaatagtg gacccagcca cgctcagggt gtgcatgcct tcaggcccaa agatgaacag
     2881 gccgggaaac atcagccaac tagaatagta cacggcagtc atggccagca ccagtttctt
     2941 acagccgccg gcaggcatga tatagtaaga ctccacgtag atgattccag cgttgtagaa
     3001 tgtctgggtt ccatacacca ggccgataca gtaaaacagc cacttcaccc acccagtggc
     3061 cagagcggct gtcactccca tgcaaatagt tcccaggtcg gacaccagca gagccattgt
     3121 ccgcttattg tatgcctcac tcaggccggt gatgttagac agatgaatca ggatcacggg
     3181 acatgtcagc agccattcgg catatctcag ccatggggta atgttccctc cgtacaggaa
     3241 cagcatagca ggactggtga actcgaaata gatctccaga atcactttga tcagctccac
     3301 ggagcacacg tacacttcct cccatcccac gctggcctta taggcgtgcc agccgtagaa
     3361 agccagaaac agggcagaca ggaacacggc aaaccactgg aagcccctag ctgtttcctg
     3421 ctcccacagg gccccatagt agatgtagat ggagctgcac tgttcctcat acccatgcac
     3481 ggggtggtag ggtggattca cttgatatcc agctggggca tggtactcac tgcctccggc
     3541 agaatcgttt ccaggtgggg cgctgtgagc gtctggagca ggagctccgc ccatggtggc
     3601 ggagctagct ggcaattcgt gagcgtcggt tgaacagcga agttcctata ctttctagag
     3661 aataggaact tcggaatagg aacttccgtt gggattcttc ctattttgat ccaagcatca
     3721 ccatcgaccc tctagtccag atctcaccat cgacccgaag ttcctatacc ttttgaagaa
     3781 taggaacttc ggaataggaa cttcaatccc cgcaataagc tcccatcaag cttatcgata
     3841 atcaacctct ggattacaaa atttgtgaaa gattgactgg tattcttaac tatgttgctc
     3901 cttttacgct atgtggatac gctgctttaa tgcctttgta tcatgctatt gcttcccgta
     3961 tggctttcat tttctcctcc ttgtataaat cctggttgct gtctctttat gaggagttgt
     4021 ggcccgttgt caggcaacgt ggcgtggtgt gcactgtgtt tgctgacgca acccccactg
     4081 gttggggcat tgccaccacc tgtcagctcc tttccgggac tttcgctttc cccctcccta
     4141 ttgccacggc ggaactcatc gccgcctgcc ttgcccgctg ctggacaggg gctcggctgt
     4201 tgggcactga caattccgtg gtgttgtcgg ggaaatcatc gtcctttcct tggctgctcg
     4261 cctatgttgc cacctggatt ctgcgcggga cgtccttctg ctacgtccct tcggccctca
     4321 atccagcgga ccttccttcc cgcggcctgc tgccggctct gcggcctctt ccgcgtcttc
     4381 gccttcgccc tcagacgagt cggatctccc tttgggccgc ctccccgcat cgataccgag
     4441 cgctgctcga gagatctacg ggtggcatcc ctgtgacccc tccccagtgc ctctcctggc
     4501 cctggaagtt gccactccag tgcccaccag ccttgtccta ataaaattaa gttgcatcat
     4561 tttgtctgac taggtgtcct tctataatat tatggggtgg aggggggtgg tatggagcaa
     4621 ggggcaagtt gggaagacaa cctgtagggc ctgcggggtc tattgggaac caagctggag
     4681 tgcagtggca caatcttggc tcactgcaat ctccgcctcc tgggttcaag cgattctcct
     4741 gcctcagcct cccgagttgt tgggattcca ggcatgcatg accaggctca gctaattttt
     4801 gtttttttgg tagagacggg gtttcaccat attggccagg ctggtctcca actcctaatc
     4861 tcaggtgatc tacccacctt ggcctcccaa attgctggga ttacaggcgt gaaccactgc
     4921 tcccttccct gtccttctga ttttgtaggt aaccacgtgc ggaccgagcg gccgcaggaa
     4981 cccctagtga tggagttggc cactccctct ctgcgcgctc gctcgctcac tgaggccggg
     5041 cgaccaaagg tcgcccgacg cccgggcttt gcccgggcgg cctcagtgag cgagcgagcg
     5101 cgcagctgcc tgcaggggcg cctgatgcgg tattttctcc ttacgcatct gtgcggtatt
     5161 tcacaccgca tacgtcaaag caaccatagt acgcgccctg tagcggcgca ttaagcgcgg
     5221 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     5281 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     5341 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     5401 ttgatttggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     5461 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     5521 accctatctc gggctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     5581 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgttta
     5641 caattttatg gtgcactctc agtacaatct gctctgatgc cgcatagtta agccagcccc
     5701 gacacccgcc aacacccgct gacgcgccct gacgggcttg tctgctcccg gcatccgctt
     5761 acagacaagc tgtgaccgtc tccgggagct gcatgtgtca gaggttttca ccgtcatcac
     5821 cgaaacgcgc gagacgaaag ggcctcgtga tacgcctatt tttataggtt aatgtcatga
     5881 taataatggt ttcttagacg tcaggtggca cttttcgggg aaatgtgcgc ggaaccccta
     5941 tttgtttatt tttctaaata cattcaaata tgtatccgct catgagacaa taaccctgat
     6001 aaatgcttca ataatattga aaaaggaaga gtatgagtat tcaacatttc cgtgtcgccc
     6061 ttattccctt ttttgcggca ttttgccttc ctgtttttgc tcacccagaa acgctggtga
     6121 aagtaaaaga tgctgaagat cagttgggtg cacgagtggg ttacatcgaa ctggatctca
     6181 acagcggtaa gatccttgag agttttcgcc ccgaagaacg ttttccaatg atgagcactt
     6241 ttaaagttct gctatgtggc gcggtattat cccgtattga cgccgggcaa gagcaactcg
     6301 gtcgccgcat acactattct cagaatgact tggttgagta ctcaccagtc acagaaaagc
     6361 atcttacgga tggcatgaca gtaagagaat tatgcagtgc tgccataacc atgagtgata
     6421 acactgcggc caacttactt ctgacaacga tcggaggacc gaaggagcta accgcttttt
     6481 tgcacaacat gggggatcat gtaactcgcc ttgatcgttg ggaaccggag ctgaatgaag
     6541 ccataccaaa cgacgagcgt gacaccacga tgcctgtagc aatggcaaca acgttgcgca
     6601 aactattaac tggcgaacta cttactctag cttcccggca acaattaata gactggatgg
     6661 aggcggataa agttgcagga ccacttctgc gctcggccct tccggctggc tggtttattg
     6721 ctgataaatc tggagccggt gagcgtgggt ctcgcggtat cattgcagca ctggggccag
     6781 atggtaagcc ctcccgtatc gtagttatct acacgacggg gagtcaggca actatggatg
     6841 aacgaaatag acagatcgct gagataggtg cctcactgat taagcattgg taactgtcag
     6901 accaagttta ctcatatata ctttagattg atttaaaact tcatttttaa tttaaaagga
     6961 tctaggtgaa gatccttttt gataatctca tgaccaaaat cccttaacgt gagttttcgt
     7021 tccactgagc gtcagacccc gtagaaaaga tcaaaggatc ttcttgagat cctttttttc
     7081 tgcgcgtaat ctgctgcttg caaacaaaaa aaccaccgct accagcggtg gtttgtttgc
     7141 cggatcaaga gctaccaact ctttttccga aggtaactgg cttcagcaga gcgcagatac
     7201 caaatactgt ccttctagtg tagccgtagt taggccacca cttcaagaac tctgtagcac
     7261 cgcctacata cctcgctctg ctaatcctgt taccagtggc tgctgccagt ggcgataagt
     7321 cgtgtcttac cgggttggac tcaagacgat agttaccgga taaggcgcag cggtcgggct
     7381 gaacgggggg ttcgtgcaca cagcccagct tggagcgaac gacctacacc gaactgagat
     7441 acctacagcg tgagctatga gaaagcgcca cgcttcccga agggagaaag gcggacaggt
     7501 atccggtaag cggcagggtc ggaacaggag agcgcacgag ggagcttcca gggggaaacg
     7561 cctggtatct ttatagtcct gtcgggtttc gccacctctg acttgagcgt cgatttttgt
     7621 gatgctcgtc aggggggcgg agcctatgga aaaacgccag caacgcggcc tttttacggt
     7681 tcctggcctt ttgctggcct tttgctcaca tgt
//
