LOCUS       sequence-448714-        6524 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cation channelrhodopsin ChrimsonR fused to mRuby2 fluorophore and 
            targeted to the neuronal soma and proximal dendrites in a viral 
            vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 6524)
  AUTHORS   Pegard NC, Mardinly AR, Oldenburg IA, Sridharan S, Waller L, Adesnik
            H
  TITLE     Three-dimensional scanless holographic optogenetics with temporal 
            focusing (3D-SHOT).
  JOURNAL   Nat Commun. 2017 Oct 31;8(1):1228. doi: 10.1038/s41467-017-01031-3.
  PUBMED    29089483
REFERENCE   2  (bases 1 to 6524)
  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: "Nat 
            Commun."; date: "2017-10-31"; volume: "8(1):1228. doi"; pages: " 
            10.1038/s41467-017-01031-3"
COMMENT     Sequence Label: pAAV-hSyn-DIO-ChrimsonR-mRuby2-ST
FEATURES             Location/Qualifiers
     source          1..6524
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   194..323
                     /label=AAV2 ITR
     polyA_signal    485..606
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(522..541)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     576..595
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     635..651
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     636..652
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    655..1243
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             761..772
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1170..1190
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    complement(1268..1301)
                     /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(1352..1385)
                     /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)."
     CDS             1395..2444
                     /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             2457..3167
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby, with improved photophysical properties (Lam et al., 
                     2012)"
                     /label=mRuby2
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGNPYMGTQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPKGIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVMQDTSLEDGCLVYHVQVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTH
                     MALKVDGGGHLSCSFVTTYRSKKTVGNIKMPGIHAVDHRLERLEESDNEMFVVQREHAV
                     AKFAGLGGGMDELYK"
     protein_bind    3383..3416
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    3467..3500
                     /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(3546..3993)
                     /label=hSyn promoter
                     /note="human synapsin I promoter; confers neuron-specific 
                     expression (Kugler et al., 2003)"
     repeat_region   4075..4204
                     /label=AAV2 ITR
     rep_origin      4276..4731
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(4363..4382)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     4573..4594
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        4757..4861
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(4862..4930,4931..5722)
                     /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(5080..5099)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      5893..6481
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     6382..6401
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ggccttttgc tcacatgttc tttcctgcgt tatcccctga ttctgtggat aaccgtatta
       61 ccgcctttga gtgagctgat accgctcgcc gcagccgaac gaccgagcgc agcgagtcag
      121 tgagcgagga agcggaagag cgcccaatac gcaaaccgcc tctccccgcg cgttggccga
      181 ttcattaatg cagctgcgcg ctcgctcgct cactgaggcc gcccgggcaa agcccgggcg
      241 tcgggcgacc tttggtcgcc cggcctcagt gagcgagcga gcgcgcagag agggagtggc
      301 caactccatc actaggggtt ccttgtagtt aatgattaac ccgccatgct acttatctac
      361 gtagccatgc tctaggaaga tccttatcga ttttaccaca tttgtagagg ttttacttgc
      421 tttaaaaaac ctcccacatc tccccctgaa cctgaaacat aaaatgaatg caattgttgt
      481 tgttaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
      541 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
      601 atcttatcat gtctgctcga agcggccgcc cgggtcgagg tcgacggtat cgatgcgggg
      661 aggcggccca aagggagatc cgactcgtct gagggcgaag gcgaagacgc ggaagaggcc
      721 gcagagccgg cagcaggccg cgggaaggaa ggtccgctgg attgagggcc gaagggacgt
      781 agcagaagga cgtcccgcgc agaatccagg tggcaacaca ggcgagcagc caaggaaagg
      841 acgatgattt ccccgacaac accacggaat tgtcagtgcc caacagccga gcccctgtcc
      901 agcagcgggc aaggcaggcg gcgatgagtt ccgccgtggc aatagggagg gggaaagcga
      961 aagtcccgga aaggagctga caggtggtgg caatgcccca accagtgggg gttgcgtcag
     1021 caaacacagt gcacaccacg ccacgttgcc tgacaacggg ccacaactcc tcataaagag
     1081 acagcaacca ggatttatac aaggaggaga aaatgaaagc catacgggaa gcaatagcat
     1141 gatacaaagg cattaaagca gcgtatccac atagcgtaaa aggagcaaca tagttaagaa
     1201 taccagtcaa tctttcacaa attttgtaat ccagaggttg attatcgata agcttgatat
     1261 cgaattcata acttcgtata gcatacatta tacgaagtta ttctttgcac cattctaaag
     1321 aataacagtg ataatttctg ggttaaggca aataacttcg tataggatac tttatacgaa
     1381 gttatgctag caccatggct gagctgatca gcagcgccac cagatctctg tttgccgccg
     1441 gaggcatcaa cccttggcct aacccctacc accacgagga catgggctgt ggaggaatga
     1501 cacctacagg cgagtgcttc agcaccgagt ggtggtgtga cccttcttac ggactgagcg
     1561 acgccggata cggatattgc ttcgtggagg ccacaggcgg ctacctggtc gtgggagtgg
     1621 agaagaagca ggcttggctg cacagcagag gcacaccagg agaaaagatc ggcgcccagg
     1681 tctgccagtg gattgctttc agcatcgcca tcgccctgct gacattctac ggcttcagcg
     1741 cctggaaggc cacttgcggt tgggaggagg tctacgtctg ttgcgtcgag gtgctgttcg
     1801 tgaccctgga gatcttcaag gagttcagca gccccgccac agtgtacctg tctaccggca
     1861 accacgccta ttgcctgcgc tacttcgagt ggctgctgtc ttgccccgtg atcctgatca
     1921 gactgagcaa cctgagcggc ctgaagaacg actacagcaa gcggaccatg ggcctgatcg
     1981 tgtcttgcgt gggaatgatc gtgttcggca tggccgcagg actggctacc gattggctca
     2041 agtggctgct gtatatcgtg tcttgcatct acggcggcta catgtacttc caggccgcca
     2101 agtgctacgt ggaagccaac cacagcgtgc ctaaaggcca ttgccgcatg gtcgtgaagc
     2161 tgatggccta cgcttacttc gcctcttggg gcagctaccc aatcctctgg gcagtgggac
     2221 cagaaggact gctgaagctg agcccttacg ccaacagcat cggccacagc atctgcgaca
     2281 tcatcgccaa ggagttttgg accttcctgg cccaccacct gaggatcaag atccacgagc
     2341 acatcctgat ccacggcgac atccggaaga ccaccaagat ggagatcgga ggcgaggagg
     2401 tggaagtgga agagttcgtg gaggaggagg acgaggacac agtggcggca ccggtaatgg
     2461 tgtctaaggg cgaagagctg atcaaggaaa atatgcgtat gaaggtggtc atggaaggtt
     2521 cggtcaacgg ccaccaattc aaatgcacag gtgaaggaga aggcaatccg tacatgggaa
     2581 ctcaaaccat gaggatcaaa gtcatcgagg gaggacccct gccatttgcc tttgacattc
     2641 ttgccacgtc gttcatgtat ggcagccgta cttttatcaa gtacccgaaa ggcattcctg
     2701 atttctttaa acagtccttt cctgagggtt ttacttggga aagagttacg agatacgaag
     2761 atggtggagt cgtcaccgtc atgcaggaca ccagccttga ggatggctgt ctcgtttacc
     2821 acgtccaagt cagaggggta aactttccct ccaatggtcc cgtgatgcag aagaagacca
     2881 agggttggga gcctaataca gagatgatgt atccagcaga tggtggtctg aggggataca
     2941 ctcatatggc actgaaagtt gatggtggtg gccatctgtc ttgctctttc gtaacaactt
     3001 acaggtcaaa aaagaccgtc gggaacatca agatgcccgg tatccatgcc gttgatcacc
     3061 gcctggaaag gttagaggaa agtgacaatg aaatgttcgt agtacaacgc gaacacgcag
     3121 ttgccaagtt cgccgggctt ggtggtggga tggacgagct gtacaaggcc ctagagcagt
     3181 cccagcccat cctcaacacc aaggagatgg ccccgcagag caagcctcca gaggagctgg
     3241 agatgagcag catgcccagc cccgtggccc ctctgcccgc acgcacggag ggcgtcatcg
     3301 acatgcggag catgtccagc attgacagct tcatcagctg tgccacggac ttccctgaag
     3361 ccaccagatt ctaaggcgcg ccataacttc gtataatgta tgctatacga agttattaag
     3421 aggtttcata ttgctaatag cagctacaat ccagctacca ttctgcataa cttcgtataa
     3481 agtatcctat acgaagttat tccggagtcg actctagagg atccttgcta tcagcttgaa
     3541 ttcgactgcg ctctcaggca cgacacgact cctccgctgc ccaccgcaga ctgaggcagc
     3601 gctgagtcgc cggcgccgca gcgcagatgg tcgcgcccgt gcccccctat ctcgcgcctc
     3661 gcgtggtgcg gtccggctgg gccggcggcg gcgcggacgc gaccaaggtg gccgggaagg
     3721 ggagtttgcg ggggaccggc gagtgacgtc agcgcgcctt cagtgctgag gcggcggtgg
     3781 cgcgcgccgc caggcggggg cgaaggcact gtccgcggtg ctgaagctgg cagtgcgcac
     3841 gcgcctcgcc gcatcctgtt tcccctcccc ctctctgata ggggatgcgc aatttgggga
     3901 atgggggttg ggtgcttgtc cagtgggtcg gggtcggtcg tcaggtaggc acccccaccc
     3961 cgcctcatcc tggtcctaaa acccacttgc actcatacgc agggccctct gcagagatct
     4021 tcctagagca tggctacgta gataagtagc atggcgggtt aatcattaac tacaaggaac
     4081 ccctagtgat ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc
     4141 gaccaaaggt cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc
     4201 gcagctggcg taatagcgaa gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc
     4261 tgaatggcga atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta
     4321 cgcgcagcgt gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc
     4381 cttcctttct cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt
     4441 tagggttccg atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg
     4501 gttcacgtag tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca
     4561 cgttctttaa tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct
     4621 attcttttga tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga
     4681 tttaacaaaa atttaacgcg aattttaaca aaatattaac gcttacaatt taggtggcac
     4741 ttttcgggga aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat
     4801 gtatccgctc atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag
     4861 tatgagtatt caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc
     4921 tgtttttgct cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc
     4981 acgagtgggt tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc
     5041 cgaagaacgt tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc
     5101 ccgtattgac gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt
     5161 ggttgagtac tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt
     5221 atgcagtgct gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat
     5281 cggaggaccg aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct
     5341 tgatcgttgg gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat
     5401 gcctgtagca atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc
     5461 ttcccggcaa caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg
     5521 ctcggccctt ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc
     5581 tcgcggtatc attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta
     5641 cacgacgggg agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc
     5701 ctcactgatt aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga
     5761 tttaaaactt catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat
     5821 gaccaaaatc ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat
     5881 caaaggatct tcttgagatc ctttttttct gcgcgtaatc tgctgcttgc aaacaaaaaa
     5941 accaccgcta ccagcggtgg tttgtttgcc ggatcaagag ctaccaactc tttttccgaa
     6001 ggtaactggc ttcagcagag cgcagatacc aaatactgtt cttctagtgt agccgtagtt
     6061 aggccaccac ttcaagaact ctgtagcacc gcctacatac ctcgctctgc taatcctgtt
     6121 accagtggct gctgccagtg gcgataagtc gtgtcttacc gggttggact caagacgata
     6181 gttaccggat aaggcgcagc ggtcgggctg aacggggggt tcgtgcacac agcccagctt
     6241 ggagcgaacg acctacaccg aactgagata cctacagcgt gagctatgag aaagcgccac
     6301 gcttcccgaa gggagaaagg cggacaggta tccggtaagc ggcagggtcg gaacaggaga
     6361 gcgcacgagg gagcttccag ggggaaacgc ctggtatctt tatagtcctg tcgggtttcg
     6421 ccacctctga cttgagcgtc gatttttgtg atgctcgtca ggggggcgga gcctatggaa
     6481 aaacgccagc aacgcggcct ttttacggtt cctggccttt tgct
//
