LOCUS       sequence-176952-        6469 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  AAV-mediated expression of ChrimsonR-GFP under the Syn promoter, in 
            floxed/reversed (Cre-dependent) manner. Using bGHpA signal..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6469)
  AUTHORS   Klapoetke NC, Murata Y, Kim SS, Pulver SR, Birdsey-Benson A, Cho YK,
            Morimoto TK, Chuong AS, Carpenter EJ, Tian Z, Wang J, Xie Y, Yan Z, 
            Zhang Y, Chow BY, Surek B, Melkonian M, Jayaraman V, 
            Constantine-Paton M, Wong GK, Boyden ES
  TITLE     Independent optical excitation of distinct neural populations.
  JOURNAL   Nat Methods. 2014 Mar;11(3):338-46. doi: 10.1038/nmeth.2836. Epub 
            2014 Feb 9.
  PUBMED    24509633
REFERENCE   2  (bases 1 to 6469)
  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/nmeth.2836"; journalName: "Nat Methods"; date: "2014-03"; 
            volume: "11"; issue: "3"; pages: "338-46"
COMMENT     Sequence Label: pAAV-Syn-FLEX-rc [ChrimsonR-GFP]
FEATURES             Location/Qualifiers
     source          1..6469
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   477..606
                     /label=AAV2 ITR
     rep_origin      678..1133
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(765..784)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     975..996
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        1159..1263
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(1264..1332,1333..2124)
                     /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(1482..1501)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      2295..2883
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     2784..2803
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   3120..3249
                     /label=AAV2 ITR
     polyA_signal    3419..3626
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     misc_feature    3667..4255
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             3773..3784
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     4182..4202
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    4288..4321
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    4392..4425
                     /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      4447..4456
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             4453..5502
                     /codon_start=1
                     /product="red light-drivable channelrhodopsin from 
                     Chlamydomonas noctigama (Klapoetke et al., 2014)"
                     /label=ChrimsonR
                     /note="K176R mutant with accelerated closing kinetics"
                     /translation="MAELISSATRSLFAAGGINPWPNPYHHEDMGCGGMTPTGECFSTE
                     WWCDPSYGLSDAGYGYCFVEATGGYLVVGVEKKQAWLHSRGTPGEKIGAQVCQWIAFSI
                     AIALLTFYGFSAWKATCGWEEVYVCCVEVLFVTLEIFKEFSSPATVYLSTGNHAYCLRY
                     FEWLLSCPVILIRLSNLSGLKNDYSKRTMGLIVSCVGMIVFGMAAGLATDWLKWLLYIV
                     SCIYGGYMYFQAAKCYVEANHSVPKGHCRMVVKLMAYAYFASWGSYPILWAVGPEGLLK
                     LSPYANSIGHSICDIIAKEFWTFLAHHLRIKIHEHILIHGDIRKTTKMEIGGEEVEVEE
                     FVEEEDEDTV"
     CDS             join(5521..5523,5524..6237)
                     /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(5521..5523,5524..6234)
                     /codon_start=1
                     /product="enhanced GFP"
                     /label=EGFP
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTLTYGVQCFSRYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSTQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(5563..5584)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5824..5843)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6171..6192
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     protein_bind    complement(6251..6284)
                     /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(6347..6380)
                     /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)."
     promoter        complement(join(6396..6469,1..374))
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
ORIGIN
        1 agcgcagatg gtcgcgcccg tgccccccta tctcgcgcct cgcgtggtgc ggtccggctg
       61 ggccggcggc ggcgcggacg cgaccaaggt ggccgggaag gggagtttgc gggggaccgg
      121 cgagtgacgt cagcgcgcct tcagtgctga ggcggcggtg gcgcgcgccg ccaggcgggg
      181 gcgaaggcac tgtccgcggt gctgaagctg gcagtgcgca cgcgcctcgc cgcatcctgt
      241 ttcccctccc cctctctgat aggggatgcg caatttgggg aatgggggtt gggtgcttgt
      301 ccagtgggtc ggggtcggtc gtcaggtagg cacccccacc ccgcctcatc ctggtcctaa
      361 aacccacttg cactcatacg cagggccctc tgcagtctag acacttaatt aacctacgat
      421 cttcctagag catggctacg tagataagta gcatggcggg ttaatcatta actacaagga
      481 acccctagtg atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg
      541 gcgaccaaag gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc
      601 gcgcagctgg cgtaatagcg aagaggcccg caccgatcgc ccttcccaac agttgcgcag
      661 cctgaatggc gaatgggacg cgccctgtag cggcgcatta agcgcggcgg gtgtggtggt
      721 tacgcgcagc gtgaccgcta cacttgccag cgccctagcg cccgctcctt tcgctttctt
      781 cccttccttt ctcgccacgt tcgccggctt tccccgtcaa gctctaaatc gggggctccc
      841 tttagggttc cgatttagtg ctttacggca cctcgacccc aaaaaacttg attagggtga
      901 tggttcacgt agtgggccat cgccctgata gacggttttt cgccctttga cgttggagtc
      961 cacgttcttt aatagtggac tcttgttcca aactggaaca acactcaacc ctatctcggt
     1021 ctattctttt gatttataag ggattttgcc gatttcggcc tattggttaa aaaatgagct
     1081 gatttaacaa aaatttaacg cgaattttaa caaaatatta acgcttacaa tttaggtggc
     1141 acttttcggg gaaatgtgcg cggaacccct atttgtttat ttttctaaat acattcaaat
     1201 atgtatccgc tcatgagaca ataaccctga taaatgcttc aataatattg aaaaaggaag
     1261 agtatgagta ttcaacattt ccgtgtcgcc cttattccct tttttgcggc attttgcctt
     1321 cctgtttttg ctcacccaga aacgctggtg aaagtaaaag atgctgaaga tcagttgggt
     1381 gcacgagtgg gttacatcga actggatctc aacagcggta agatccttga gagttttcgc
     1441 cccgaagaac gttttccaat gatgagcact tttaaagttc tgctatgtgg cgcggtatta
     1501 tcccgtattg acgccgggca agagcaactc ggtcgccgca tacactattc tcagaatgac
     1561 ttggttgagt actcaccagt cacagaaaag catcttacgg atggcatgac agtaagagaa
     1621 ttatgcagtg ctgccataac catgagtgat aacactgcgg ccaacttact tctgacaacg
     1681 atcggaggac cgaaggagct aaccgctttt ttgcacaaca tgggggatca tgtaactcgc
     1741 cttgatcgtt gggaaccgga gctgaatgaa gccataccaa acgacgagcg tgacaccacg
     1801 atgcctgtag caatggcaac aacgttgcgc aaactattaa ctggcgaact acttactcta
     1861 gcttcccggc aacaattaat agactggatg gaggcggata aagttgcagg accacttctg
     1921 cgctcggccc ttccggctgg ctggtttatt gctgataaat ctggagccgg tgagcgtggg
     1981 tctcgcggta tcattgcagc actggggcca gatggtaagc cctcccgtat cgtagttatc
     2041 tacacgacgg ggagtcaggc aactatggat gaacgaaata gacagatcgc tgagataggt
     2101 gcctcactga ttaagcattg gtaactgtca gaccaagttt actcatatat actttagatt
     2161 gatttaaaac ttcattttta atttaaaagg atctaggtga agatcctttt tgataatctc
     2221 atgaccaaaa tcccttaacg tgagttttcg ttccactgag cgtcagaccc cgtagaaaag
     2281 atcaaaggat cttcttgaga tccttttttt ctgcgcgtaa tctgctgctt gcaaacaaaa
     2341 aaaccaccgc taccagcggt ggtttgtttg ccggatcaag agctaccaac tctttttccg
     2401 aaggtaactg gcttcagcag agcgcagata ccaaatactg ttcttctagt gtagccgtag
     2461 ttaggccacc acttcaagaa ctctgtagca ccgcctacat acctcgctct gctaatcctg
     2521 ttaccagtgg ctgctgccag tggcgataag tcgtgtctta ccgggttgga ctcaagacga
     2581 tagttaccgg ataaggcgca gcggtcgggc tgaacggggg gttcgtgcac acagcccagc
     2641 ttggagcgaa cgacctacac cgaactgaga tacctacagc gtgagctatg agaaagcgcc
     2701 acgcttcccg aagggagaaa ggcggacagg tatccggtaa gcggcagggt cggaacagga
     2761 gagcgcacga gggagcttcc agggggaaac gcctggtatc tttatagtcc tgtcgggttt
     2821 cgccacctct gacttgagcg tcgatttttg tgatgctcgt caggggggcg gagcctatgg
     2881 aaaaacgcca gcaacgcggc ctttttacgg ttcctggcct tttgctggcc ttttgctcac
     2941 atgttctttc ctgcgttatc ccctgattct gtggataacc gtattaccgc ctttgagtga
     3001 gctgataccg ctcgccgcag ccgaacgacc gagcgcagcg agtcagtgag cgaggaagcg
     3061 gaagagcgcc caatacgcaa accgcctctc cccgcgcgtt ggccgattca ttaatgcagc
     3121 tgcgcgctcg ctcgctcact gaggccgccc gggcaaagcc cgggcgtcgg gcgacctttg
     3181 gtcgcccggc ctcagtgagc gagcgagcgc gcagagaggg agtggccaac tccatcacta
     3241 ggggttcctt gtagttaatg attaacccgc catgctactt atctacgtag ccatgctcta
     3301 ggaagatcct tatcgatttt accacatttg tagaggtttt acttgcttta aaaaacctcc
     3361 cacatctccc cctgaacctg aaacataaaa tgaatgcaat tgtacagtcc ggaccgcatc
     3421 cccagcatgc ctgctattgt cttcccaatc ctcccccttg ctgtcctgcc ccaccccacc
     3481 ccccagaata gaatgacacc tactcagaca atgcgatgca atttcctcat tttattagga
     3541 aaggacagtg ggagtggcac cttccagggt caaggaaggc acgggggagg ggcaaacaac
     3601 agatggctgg caactagaag gcacagtcga gctttatgag cggccgcccg ggtcgacggt
     3661 atcgatgcgg ggaggcggcc caaagggaga tccgactcgt ctgagggcga aggcgaagac
     3721 gcggaagagg ccgcagagcc ggcagcaggc cgcgggaagg aaggtccgct ggattgaggg
     3781 ccgaagggac gtagcagaag gacgtcccgc gcagaatcca ggtggcaaca caggcgagca
     3841 gccaaggaaa ggacgatgat ttccccgaca acaccacgga attgtcagtg cccaacagcc
     3901 gagcccctgt ccagcagcgg gcaaggcagg cggcgatgag ttccgccgtg gcaataggga
     3961 gggggaaagc gaaagtcccg gaaaggagct gacaggtggt ggcaatgccc caaccagtgg
     4021 gggttgcgtc agcaaacaca gtgcacacca cgccacgttg cctgacaacg ggccacaact
     4081 cctcataaag agacagcaac caggatttat acaaggagga gaaaatgaaa gccatacggg
     4141 aagcaatagc atgatacaaa ggcattaaag cagcgtatcc acatagcgta aaaggagcaa
     4201 catagttaag aataccagtc aatctttcac aaattttgta atccagaggt tgattatcga
     4261 taagcttcgc gaacctcttc gagggacata acttcgtata atgtatgcta tacgaagtta
     4321 tgggtcgatg gtgagatctg gactagaggg tcgatggtga tgcttggatc aaaataggaa
     4381 gaatcccaat gataacttcg tataaagtat cctatacgaa gttatactag gtacctaggc
     4441 cgcgccgcca ccatggctga gctgatcagc agcgccacca gatctctgtt tgccgccgga
     4501 ggcatcaacc cttggcctaa cccctaccac cacgaggaca tgggctgtgg aggaatgaca
     4561 cctacaggcg agtgcttcag caccgagtgg tggtgtgacc cttcttacgg actgagcgac
     4621 gccggatacg gatattgctt cgtggaggcc acaggcggct acctggtcgt gggagtggag
     4681 aagaagcagg cttggctgca cagcagaggc acaccaggag aaaagatcgg cgcccaggtc
     4741 tgccagtgga ttgctttcag catcgccatc gccctgctga cattctacgg cttcagcgcc
     4801 tggaaggcca cttgcggttg ggaggaggtc tacgtctgtt gcgtcgaggt gctgttcgtg
     4861 accctggaga tcttcaagga gttcagcagc cccgccacag tgtacctgtc taccggcaac
     4921 cacgcctatt gcctgcgcta cttcgagtgg ctgctgtctt gccccgtgat cctgatcaga
     4981 ctgagcaacc tgagcggcct gaagaacgac tacagcaagc ggaccatggg cctgatcgtg
     5041 tcttgcgtgg gaatgatcgt gttcggcatg gccgcaggac tggctaccga ttggctcaag
     5101 tggctgctgt atatcgtgtc ttgcatctac ggcggctaca tgtacttcca ggccgccaag
     5161 tgctacgtgg aagccaacca cagcgtgcct aaaggccatt gccgcatggt cgtgaagctg
     5221 atggcctacg cttacttcgc ctcttggggc agctacccaa tcctctgggc agtgggacca
     5281 gaaggactgc tgaagctgag cccttacgcc aacagcatcg gccacagcat ctgcgacatc
     5341 atcgccaagg agttttggac cttcctggcc caccacctga ggatcaagat ccacgagcac
     5401 atcctgatcc acggcgacat ccggaagacc accaagatgg agatcggagg cgaggaggtg
     5461 gaagtggaag agttcgtgga ggaggaggac gaggacacag tggcggcacc ggtagtagca
     5521 gtgagcaagg gcgaggagct gttcaccggg gtggtgccca tcctggtcga gctggacggc
     5581 gacgtaaacg gccacaagtt cagcgtgtcc ggcgagggcg agggcgatgc cacctacggc
     5641 aagctgaccc tgaagttcat ttgcaccacc ggcaagctgc ccgtgccctg gcccaccctc
     5701 gtgaccaccc tgacctacgg cgtgcagtgc ttcagccgct accccgacca catgaagcag
     5761 cacgacttct tcaagtccgc catgcccgaa ggctacgtcc aggagcgcac catcttcttc
     5821 aaggacgacg gcaactacaa gacccgcgcc gaggtgaagt tcgagggcga caccctggtg
     5881 aaccgcatcg agctgaaggg catcgacttc aaggaggacg gcaacatcct ggggcacaag
     5941 ctggagtaca actacaacag ccacaacgtc tatatcatgg ccgacaagca gaagaacggc
     6001 atcaaggtga acttcaagat ccgccacaac atcgaggacg gcagcgtgca gctcgccgac
     6061 cactaccagc agaacacccc catcggcgac ggccccgtgc tgctgcccga caaccactac
     6121 ctgagcaccc agtccgccct gagcaaagac cccaacgaga agcgcgatca catggtcctg
     6181 ctggagttcg tgaccgccgc cgggatcact ctcggcatgg acgagctgta caagtaatga
     6241 gctagctagt ataacttcgt atagcataca ttatacgaag ttatgggtcg atggtgatgc
     6301 ttggcaattc gggtcgatgg tgaagcattg gtcttcctat tttgatataa cttcgtatag
     6361 gatactttat acgaagttat gggaattctt ctcgactgcg ctctcaggca cgacacgact
     6421 cctccgctgc ccaccgcaga ctgaggcagc gctgagtcgc cggcgccgc
//
