LOCUS       sequence-244666-        6706 bp    DNA     linear   UNA 15-DEC-2025
DEFINITION  For AAV based expression of CheRiff in Cre-Positive cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      natural DNA sequence
  ORGANISM  unspecified
REFERENCE   1  (bases 1 to 6706)
  AUTHORS   Farhi SL, Parot VJ, Grama A, Yamagata M, Abdelfattah AS, Adam Y, Lou
            S, Jun Kim J, Campbell RE, Cox DD, Cohen AE
  TITLE     Wide-area all-optical neurophysiology in acute brain slices.
  JOURNAL   J Neurosci. 2019 Apr 5. pii: JNEUROSCI.0168-19.2019. doi: 
            10.1523/JNEUROSCI.0168-19.2019.
  PUBMED    30952812
REFERENCE   2  (bases 1 to 6706)
  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: "J Neurosci.
            2019 Apr 5. pii: JNEUROSCI.0168-19.2019. doi: 
            10.1523/JNEUROSCI.0168-19.2019."
COMMENT     Sequence Label: SL016: AAV-CAG-FLEX-CheRiff-GFP
FEATURES             Location/Qualifiers
     source          1..6706
                     /mol_type="genomic DNA"
                     /organism="unspecified"
     rep_origin      complement(8..463)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(145..166)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     357..376
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     repeat_region   535..664
                     /label=AAV2 ITR
     polyA_signal    826..947
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(863..882)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     917..936
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     misc_feature    990..1578
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1096..1107
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1505..1525
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    1669..1702
                     /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)."
     regulatory      1796..1805
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1802..2749
                     /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"
     CDS             join(2819..2821,2822..3535)
                     /codon_start=1
                     /product="the original enhanced GFP (Yang et al., 1996)"
                     /label=EGFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     CDS             join(2819..2821,2822..3532)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(2861..2882)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(3122..3141)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     3469..3490
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(3578..3611)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    complement(3674..3707)
                     /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)."
     primer_bind     complement(3742..3761)
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     complement(3823..3845)
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     promoter        complement(4124..4401)
                     /label=chicken beta-actin promoter
     rep_origin      complement(4904..5492)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(4984..5003)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5663..6454,6455..6523))
                     /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     6286..6305
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6524..6628)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 gccacctaaa ttgtaagcgt taatattttg ttaaaattcg cgttaaattt ttgttaaatc
       61 agctcatttt ttaaccaata ggccgaaatc ggcaaaatcc cttataaatc aaaagaatag
      121 accgagatag ggttgagtgt tgttccagtt tggaacaaga gtccactatt aaagaacgtg
      181 gactccaacg tcaaagggcg aaaaaccgtc tatcagggcg atggcccact acgtgaacca
      241 tcaccctaat caagtttttt ggggtcgagg tgccgtaaag cactaaatcg gaaccctaaa
      301 gggagccccc gatttagagc ttgacgggga aagccggcga acgtggcgag aaaggaaggg
      361 aagaaagcga aaggagcggg cgctagggcg ctggcaagtg tagcggtcac gctgcgcgta
      421 accaccacac ccgccgcgct taatgcgccg ctacagggcg cgtcccattc gccattcagg
      481 ctgcgcaact gttgggaagg gcgatcggtg cgggcctctt cgctattacg ccagctgcgc
      541 gctcgctcgc tcactgaggc cgcccgggca aagcccgggc gtcgggcgac ctttggtcgc
      601 ccggcctcag tgagcgagcg agcgcgcaga gagggagtgg ccaactccat cactaggggt
      661 tccttgtagt taatgattaa cccgccatgc tacttatcta cgtagccatg ctctaggaag
      721 atccttatcg attttaccac atttgtagag gttttacttg ctttaaaaaa cctcccacat
      781 ctccccctga acctgaaaca taaaatgaat gcaattgttg ttgttaactt gtttattgca
      841 gcttataatg gttacaaata aagcaatagc atcacaaatt tcacaaataa agcatttttt
      901 tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg tatcttatca tgtctgctcg
      961 aagcggccgc ccgggtcgac ggtatcgatg cggggaggcg gcccaaaggg agatccgact
     1021 cgtctgaggg cgaaggcgaa gacgcggaag aggccgcaga gccggcagca ggccgcggga
     1081 aggaaggtcc gctggattga gggccgaagg gacgtagcag aaggacgtcc cgcgcagaat
     1141 ccaggtggca acacaggcga gcagccaagg aaaggacgat gatttccccg acaacaccac
     1201 ggaattgtca gtgcccaaca gccgagcccc tgtccagcag cgggcaaggc aggcggcgat
     1261 gagttccgcc gtggcaatag ggagggggaa agcgaaagtc ccggaaagga gctgacaggt
     1321 ggtggcaatg ccccaaccag tgggggttgc gtcagcaaac acagtgcaca ccacgccacg
     1381 ttgcctgaca acgggccaca actcctcata aagagacagc aaccaggatt tatacaagga
     1441 ggagaaaatg aaagccatac gggaagcaat agcatgatac aaaggcatta aagcagcgta
     1501 tccacatagc gtaaaaggag caacatagtt aagaatacca gtcaatcttt cacaaatttt
     1561 gtaatccaga ggttgattat cgataagctt gatatcgaag ttcgatggtg agatctggac
     1621 tagagggtcg atggtgatgc ttggatcaaa ataggaagaa tcccaatgat aacttcgtat
     1681 aaagtatcct atacgaagtt atactagaat tcggtacaat caagctggtt ctccatttgc
     1741 actcaggagc acgggcaggc gagtggcccc tagttctggg ggcagcgggg gatccgccac
     1801 catgggcgga gctcctgctc cagacgctca cagcgcccca cctggaaacg attctgccgg
     1861 aggcagtgag taccatgccc cagctggata tcaagtgaat ccaccctacc accccgtgca
     1921 tgggtatgag gaacagtgca gctccatcta catctactat ggggccctgt gggagcagga
     1981 aacagctagg ggcttccagt ggtttgccgt gttcctgtct gccctgtttc tggctttcta
     2041 cggctggcac gcctataagg ccagcgtggg atgggaggaa gtgtacgtgt gctccgtgga
     2101 gctgatcaaa gtgattctgg agatctattt cgagttcacc agtcctgcta tgctgttcct
     2161 gtacggaggg aacattaccc catggctgag atatgccgaa tggctgctga catgtcccgt
     2221 gatcctgatt catctgtcta acatcaccgg cctgagtgag gcatacaata agcggacaat
     2281 ggctctgctg gtgtccgacc tgggaactat ttgcatggga gtgacagccg ctctggccac
     2341 tgggtgggtg aagtggctgt tttactgtat cggcctggtg tatggaaccc agacattcta
     2401 caacgctgga atcatctacg tggagtctta ctatatcatg cctgccggcg gctgtaagaa
     2461 actggtgctg gccatgactg ccgtgtacta ttctagttgg ctgatgtttc ccggcctgtt
     2521 catctttggg cctgaaggca tgcacaccct gagcgtggct gggtccacta ttggccatac
     2581 catcgccgac ctgctgtcca agaatatttg gggactgctg gggcacttcc tgcggatcaa
     2641 aattcacgag catatcatta tgtacggcga tatcaggaga ccagtgagct cccagtttct
     2701 gggacgcaag gtggacgtgc tggccttcgt gacagaggaa gataaagtgg cggccgccaa
     2761 gagcaggatc accagcgagg gcgagtacat ccccctggac cagatcgaca tcaacgtggt
     2821 gagcaagggc gaggagctgt tcaccggggt ggtgcccatc ctggtcgagc tggacggcga
     2881 cgtaaacggc cacaagttca gcgtgtccgg cgagggcgag ggcgatgcca cctacggcaa
     2941 gctgaccctg aagttcattt gcaccaccgg caagctgccc gtgccctggc ccaccctcgt
     3001 gaccaccctg acctacggcg tgcagtgctt cagccgctac cccgaccaca tgaagcagca
     3061 cgacttcttc aagtccgcca tgcccgaagg ctacgtccag gagcgcacca tcttcttcaa
     3121 ggacgacggc aactacaaga cccgcgccga ggtgaagttc gagggcgaca ccctggtgaa
     3181 ccgcatcgag ctgaagggca tcgacttcaa ggaggacggc aacatcctgg ggcacaagct
     3241 ggagtacaac tacaacagcc acaacgtcta tatcatggcc gacaagcaga agaacggcat
     3301 caaggtgaac ttcaagatcc gccacaacat cgaggacggc agcgtgcagc tcgccgacca
     3361 ctaccagcag aacaccccca tcggcgacgg ccccgtgctg ctgcccgaca accactacct
     3421 gagcacccag tccgccctga gcaaagaccc caacgagaag cgcgatcaca tggtcctgct
     3481 ggagttcgtg accgccgccg ggatcactct cggcatggac gagctgtaca agtaagaatt
     3541 cgatatcaag cttatcgaaa gtaaggggat ccctagtata acttcgtata gcatacatta
     3601 tacgaagtta tgggtcgatg gtgatgcttg gcaattcggg tcgatggtga agcattggtc
     3661 ttcctatttt gatataactt cgtataggat actttatacg aagttatggg gatccaattc
     3721 tttgccaaaa tgatgagaca gcacaataac cagcacgttg cccaggagct gtaggaaaaa
     3781 gaagaaggca tgaacatggt tagcagaggc tctagagccg ccggtcacac gccagaagcc
     3841 gaaccccgcc ctgccccgtc ccccccgaag gcagccgtcc ccccgcggac agccccgagc
     3901 cgctcccccc gcacaaaggg ccctcccgga gcccctcaag gctttcacgc agccacagaa
     3961 aagaaacaag ccgtcattaa accaagcgct aattacagcc cggaggagaa gggccgtccc
     4021 gcccgctcac ctgtgggagt aacgcggtca gtcagagccg gggcgggcgg cgcgaggcgg
     4081 cggcggagcg gggcacgggg cgaaggcagc gcgcagcgac tcccgcccgc cgcgcgcttc
     4141 gctttttata gggccgccgc cgccgccgcc tcgccataaa aggaaacttt cggagcgcgc
     4201 cgctctgatt ggctgccgcc gcaccccccc gccccgcccc cccccccccc cccccccccc
     4261 cccccccccc cggcgcgcgc cccccccccc cccccgcccc catcgctgca caaaataatt
     4321 aaaaaataaa taaatacaaa attgggggtg gggagggggg ggagatgggg agagtgaagc
     4381 agaacgtggg gctcacctcg accatggtaa tagcgatgac taatacgtag atgtactgcc
     4441 aagtaggaaa gtcccataag gtcatgtact gggcataatg ccaggcgggc catttaccgt
     4501 cattgacgtc aatagggggc gtacttggca tatgatacac ttgatgtact gccaagtggg
     4561 cagtttaccg taaatactcc acccattgac gtcaatggaa agtccctatt ggcgttacta
     4621 tgggaacata cgtcattatt gacgtcaatg gtactcttcc tagagcatgg ctacgtagat
     4681 aagtagcatg gcgggttaat cattaactac aaggaacccc tagtgatgga gttggccact
     4741 ccctctctgc gcgctcgctc gctcgctgag gccgagcggt atcagctcac tcaaaggcgg
     4801 taatacggtt atccacagaa tcaggggata acgcaggaaa gaacatgtga gcaaaaggcc
     4861 agcaaaaggc caggaaccgt aaaaaggccg cgttgctggc gtttttccat aggctccgcc
     4921 cccctgacga gcatcacaaa aatcgacgct caagtcagag gtggcgaaac ccgacaggac
     4981 tataaagata ccaggcgttt ccccctggaa gctccctcgt gcgctctcct gttccgaccc
     5041 tgccgcttac cggatacctg tccgcctttc tcccttcggg aagcgtggcg ctttctcata
     5101 gctcacgctg taggtatctc agttcggtgt aggtcgttcg ctccaagctg ggctgtgtgc
     5161 acgaaccccc cgttcagccc gaccgctgcg ccttatccgg taactatcgt cttgagtcca
     5221 acccggtaag acacgactta tcgccactgg cagcagccac tggtaacagg attagcagag
     5281 cgaggtatgt aggcggtgct acagagttct tgaagtggtg gcctaactac ggctacacta
     5341 gaagaacagt atttggtatc tgcgctctgc tgaagccagt taccttcgga aaaagagttg
     5401 gtagctcttg atccggcaaa caaaccaccg ctggtagcgg tggttttttt gtttgcaagc
     5461 agcagattac gcgcagaaaa aaaggatctc aagaagatcc tttgatcttt tctacggggt
     5521 ctgacgctca gtggaacgaa aactcacgtt aagggatttt ggtcatgaga ttatcaaaaa
     5581 ggatcttcac ctagatcctt ttaaattaaa aatgaagttt taaatcaatc taaagtatat
     5641 atgagtaaac ttggtctgac agttaccaat gcttaatcag tgaggcacct atctcagcga
     5701 tctgtctatt tcgttcatcc atagttgcct gactccccgt cgtgtagata actacgatac
     5761 gggagggctt accatctggc cccagtgctg caatgatacc gcgagaccca cgctcaccgg
     5821 ctccagattt atcagcaata aaccagccag ccggaagggc cgagcgcaga agtggtcctg
     5881 caactttatc cgcctccatc cagtctatta attgttgccg ggaagctaga gtaagtagtt
     5941 cgccagttaa tagtttgcgc aacgttgttg ccattgctac aggcatcgtg gtgtcacgct
     6001 cgtcgtttgg tatggcttca ttcagctccg gttcccaacg atcaaggcga gttacatgat
     6061 cccccatgtt gtgcaaaaaa gcggttagct ccttcggtcc tccgatcgtt gtcagaagta
     6121 agttggccgc agtgttatca ctcatggtta tggcagcact gcataattct cttactgtca
     6181 tgccatccgt aagatgcttt tctgtgactg gtgagtactc aaccaagtca ttctgagaat
     6241 agtgtatgcg gcgaccgagt tgctcttgcc cggcgtcaat acgggataat accgcgccac
     6301 atagcagaac tttaaaagtg ctcatcattg gaaaacgttc ttcggggcga aaactctcaa
     6361 ggatcttacc gctgttgaga tccagttcga tgtaacccac tcgtgcaccc aactgatctt
     6421 cagcatcttt tactttcacc agcgtttctg ggtgagcaaa aacaggaagg caaaatgccg
     6481 caaaaaaggg aataagggcg acacggaaat gttgaatact catactcttc ctttttcaat
     6541 attattgaag catttatcag ggttattgtc tcatgagcgg atacatattt gaatgtattt
     6601 agaaaaataa acaaataggg gttccgcgca catttccccg aaaagtgcca cctaaattgt
     6661 aagcgttaat attttgttaa aattcgcgtt aaatttttgt taaatc
//
