LOCUS       sequence-445530-        7378 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cation channelrhodopsin ChroME2s targeted to the neuronal soma and 
            proximal dendrites and separated from nuclear mRuby3 by P2A in a 
            viral vector.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7378)
  AUTHORS   Sridharan S, Gajowa MA, Ogando MB, Jagadisan UK, Abdeladim L, 
            Sadahiro M, Bounds HA, Hendricks WD, Turney TS, Tayler I, Gopakumar 
            K, Oldenburg IA, Brohawn SG, Adesnik H
  TITLE     High-performance microbial opsins for spatially and temporally 
            precise perturbations of large neuronal networks.
  JOURNAL   Neuron. 2022 Jan 27. pii: S0896-6273(22)00008-3. doi: 
            10.1016/j.neuron.2022.01.008.
  PUBMED    35120626
REFERENCE   2  (bases 1 to 7378)
  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: "Neuron. 
            2022 Jan 27. pii: S0896-6273(22)00008-3. doi: 
            10.1016/j.neuron.2022.01.008."
COMMENT     Sequence Label: pAAV-CAG-DIO-ChroME2s-ST-P2A-H2B-mRuby3
FEATURES             Location/Qualifiers
     source          1..7378
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   52..181
                     /label=AAV2 ITR
     promoter        495..775
                     /label=chicken beta-actin promoter
     primer_bind     1054..1076
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1138..1157
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    complement(1208..1241)
                     /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(1292..1325)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1337..2047)
                     /codon_start=1
                     /product="monomeric red fluorescent protein derived from 
                     mRuby2, with improved brightness and photostability (Bajar 
                     et al., 2016)"
                     /label=mRuby3
                     /translation="MVSKGEELIKENMRMKVVMEGSVNGHQFKCTGEGEGRPYEGVQTM
                     RIKVIEGGPLPFAFDILATSFMYGSRTFIKYPADIPDFFKQSFPEGFTWERVTRYEDGG
                     VVTVTQDTSLEDGELVYNVKVRGVNFPSNGPVMQKKTKGWEPNTEMMYPADGGLRGYTD
                     IALKVDGGGHLHCNFVTTYRSKKTVGNIKMPGVHAVDHRLERIEESDNETYVVQREVAV
                     AKYSNLGGGMDELYK"
     CDS             complement(2057..2434)
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     CDS             complement(2441..2497)
                     /codon_start=1
                     /product="2A peptide from porcine teschovirus-1 
                     polyprotein"
                     /label=P2A
                     /note="Eukaryotic ribosomes fail to insert a peptide bond 
                     between the Gly and Pro residues, yielding separate 
                     polypeptides."
                     /translation="ATNFSLLKQAGDVEENPGP"
     CDS             complement(2708..2731)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     protein_bind    3725..3758
                     /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    3809..3842
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    3867..4455
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(3920..3940)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4338..4349)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4458..4474)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4459..4475)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    4504..4625
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4515..4534)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4569..4588
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   4787..4916
                     /label=AAV2 ITR
     rep_origin      complement(5153..5741)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5233..5252)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(5912..6703,6704..6772))
                     /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     6535..6554
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(6773..6877)
                     /gene="bla"
                     /label=AmpR promoter
     rep_origin      complement(6903..7358)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(7040..7061)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     7252..7271
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
ORIGIN
        1 cgcaactgtt gggaagggcg atcggtgcgg gcctcttcgc tattacgcca gctgcgcgct
       61 cgctcgctca ctgaggccgc ccgggcaaag cccgggcgtc gggcgacctt tggtcgcccg
      121 gcctcagtga gcgagcgagc gcgcagagag ggagtggcca actccatcac taggggttcc
      181 ttgtagttaa tgattaaccc gccatgctac ttatctacgt agccatgctc taggaagatc
      241 tgtaccattg acgtcaataa tgacgtatgt tcccatagta acgccaatag ggactttcca
      301 ttgacgtcaa tgggtggagt atttacggta aactgcccac ttggcagtac atcaagtgta
      361 tcatatgcca agtacgcccc ctattgacgt caatgacggt aaatggcccg cctggcatta
      421 tgcccagtac atgaccttat gggactttcc tacttggcag tacatctacg tattagtcat
      481 cgctattacc atggtcgagg tgagccccac gttctgcttc actctcccca tctccccccc
      541 ctccccaccc ccaattttgt atttatttat tttttaatta ttttgtgcag cgatgggggc
      601 gggggggggg gggggggggc gcgcgccagg cggggcgggg cggggcgagg ggcggggcgg
      661 ggcgaggcgg agaggtgcgg cggcagccaa tcagagcggc gcgctccgaa agtttccttt
      721 tatggcgagg cggcggcggc ggcggcccta taaaaagcga agcgcgcggc gggcgggagt
      781 cgctgcgcgc tgccttcgcc ccgtgccccg ctccgccgcc gcctcgcgcc gcccgccccg
      841 gctctgactg accgcgttac tcccacaggt gagcgggcgg gacggccctt ctcctccggg
      901 ctgtaattag cgcttggttt aatgacggct tgtttctttt ctgtggctgc gtgaaagcct
      961 tgaggggctc cgggagggcc ctttgtgcgg ggggagcggc tcggggctgt ccgcgggggg
     1021 acggctgcct tcggggggga cggggcaggg cggggttcgg cttctggcgt gtgaccggcg
     1081 gctctagagc ctctgctaac catgttcatg ccttcttctt tttcctacag ctcctgggca
     1141 acgtgctggt tattgtgctg tctcatcatt ttggcaaaga attggatcct ctagagtcga
     1201 ctccggaata acttcgtata ggatacttta tacgaagtta tgcagaatgg tagctggatt
     1261 gtagctgcta ttagcaatat gaaacctctt aataacttcg tatagcatac attatacgaa
     1321 gttatggcgc gcctcacttg tacagctcgt ccatgccacc accaaggttg ctgtatttgg
     1381 caactgccac ttctctttgc actacgtagg tttcattgtc actctcctcg atcctttcca
     1441 ggcggtgatc aacggcatgg acaccgggca tcttgatgtt cccgacggtc ttttttgacc
     1501 tgtaagttgt cacgaagttg cagtgcagat ggccaccacc atcaactttc agtgcgatgt
     1561 cagtgtatcc tctcagacca ccatctgctg gatacatcat ctctgtatta ggctcccaac
     1621 ccttggtctt cttctgcatc acgggaccat tggagggaaa gtttacccct ctgaccttga
     1681 cgttgtagac gagctcgcca tcctcaaggc tggtgtcctg cgtgacggtg acgactccac
     1741 catcttcgta tctcgtaact ctttcccaag taaaaccctc aggaaaggac tgtttaaaga
     1801 aatcagggat gtcggccggg tacttgataa aggtacggct gccatacatg aacgacgtgg
     1861 caagaatgtc aaaggcaaat ggcaggggtc ctccctcgat gactttgatc ctcatggttt
     1921 gcactccctc gtacggtctg ccttctcctt cacctgtgca tttgaattgg tggccgttga
     1981 ccgaaccttc catgaccacc ttcatacgca tattttcctt gatcagctct tcgcccttag
     2041 acaccatctc gagtgcctta gcgctggtgt acttggtgat ggccttagta ccctcggaca
     2101 cggcgtgctt ggccaactcc ccaggcagca gcaggcgcac ggccgtctgg atctccctgg
     2161 aggtgatggt cgagcgcttg ttgtaatgcg ccaggcggga agcctcacct gcgatgcgct
     2221 cgaaaatgtc gttcacaaac gaattcatga tgcccatggc cttggacgaa atgccggtgt
     2281 cagggtggac ctgcttcaga accttgtaca catagatgga atagctctcc ttgcggctgc
     2341 gcttgcgctt cttgccgcct ttcttctgcg ccttagtcac cgccttcttg gagccctttt
     2401 tcggggcggg agcagacttc gctggctctg gcatactagt aggtccaggg ttctcctcca
     2461 cgtctccagc ctgcttcagc aggctgaagt tagtagctcc gcttcctgcc gcgaatctgg
     2521 tggcttcagg gaagtccgtg gcacagctga tgaagctgtc aatgctggac atgctccgca
     2581 tgtcgatgac gccctccgtg cgtgcgggca gaggggccac ggggctgggc atgctgctca
     2641 tctccagctc ctctggaggc ttgctctgcg gggccatctc cttggtgttg aggatgggct
     2701 gggactgctt atcgtcatcg tctttgtaat ctaccggtgc cactcctccc tcctcctcct
     2761 cgtcgacgaa ggtctcgatc tccatgttct cgccggccac gttgatggtg gttgtcttcc
     2821 ggatgtcgcc gtggatcagg atgtgctcgt ggatcttcac tctcaggtgg tggcccagga
     2881 agccccaagt gttcttggcg atcagatcca ggatcaggtg gccgattccg ctggtgtaag
     2941 gtgtgatcag gcccagtccc tcgtgtccgg cgatgaacat cacggggaac atcagccaag
     3001 agcagaagaa gacgtaggcc atgatcttgc aaatcttccg gcagacgcct ttgggcaggg
     3061 tgtggtagct ctcgatatac accttggcga tctggaagaa gcaagtcacg ccgtagaaca
     3121 ggccgatcat gaagaacagg atcttcaggg ggccctttgt aaaggcggct gtgatccccc
     3181 acacgatgtt gccgatgtcg gtcaccagga tggtcatggt ccgcttgctg tactcttcgt
     3241 gcagtccggt caggttgctc aggtggatca ggatcacggg gcaagtcagg agccactcgc
     3301 tgtaccgcag ccaaatcacg gctcctccgt tggtctggta cactgtggct gggctgtcga
     3361 cctcgtggaa cagctcgatg aagcacttga cgagctcgat cacgcagacg tagacctcct
     3421 cccagccggt tgtggccttg aagctgtgcc aaccgtagaa gatgagctgc acgatagcca
     3481 cgatcacggt gaaccactgg agtcccactg cgatcttgtg ctgcagctcg gttccgtggt
     3541 tgatgtgggg gaagcaaccg tgatcggctc cggctgttgt ggcgttagag gtttctccgt
     3601 gagcgtctgc tgctggtgtc accactgctg cggcgctcag caggcctcta attgtggctt
     3661 cggcgctagg cacggctgag ataaaggcgt gggtcattgt ggcggctgtt tccatactgc
     3721 tagcataact tcgtataaag tatcctatac gaagttattt gccttaaccc agaaattatc
     3781 actgttattc tttagaatgg tgcaaagaat aacttcgtat aatgtatgct atacgaagtt
     3841 atgaattcga tatcaagctt atcgataatc aacctctgga ttacaaaatt tgtgaaagat
     3901 tgactggtat tcttaactat gttgctcctt ttacgctatg tggatacgct gctttaatgc
     3961 ctttgtatca tgctattgct tcccgtatgg ctttcatttt ctcctccttg tataaatcct
     4021 ggttgctgtc tctttatgag gagttgtggc ccgttgtcag gcaacgtggc gtggtgtgca
     4081 ctgtgtttgc tgacgcaacc cccactggtt ggggcattgc caccacctgt cagctccttt
     4141 ccgggacttt cgctttcccc ctccctattg ccacggcgga actcatcgcc gcctgccttg
     4201 cccgctgctg gacaggggct cggctgttgg gcactgacaa ttccgtggtg ttgtcgggga
     4261 aatcatcgtc ctttccttgg ctgctcgcct gtgttgccac ctggattctg cgcgggacgt
     4321 ccttctgcta cgtcccttcg gccctcaatc cagcggacct tccttcccgc ggcctgctgc
     4381 cggctctgcg gcctcttccg cgtcttcgcc ttcgccctca gacgagtcgg atctcccttt
     4441 gggccgcctc cccgcatcga taccgtcgac ctcgacccgg gcggccgctt cgagcagaca
     4501 tgataagata cattgatgag tttggacaaa ccacaactag aatgcagtga aaaaaatgct
     4561 ttatttgtga aatttgtgat gctattgctt tatttgtaac cattataagc tgcaataaac
     4621 aagttaacaa caacaattgc attcatttta tgtttcaggt tcagggggag atgtgggagg
     4681 ttttttaaag caagtaaaac ctctacaaat gtggtaaaat cgataaggat cttcctagag
     4741 catggctacg tagataagta gcatggcggg ttaatcatta actacaagga acccctagtg
     4801 atggagttgg ccactccctc tctgcgcgct cgctcgctca ctgaggccgg gcgaccaaag
     4861 gtcgcccgac gcccgggctt tgcccgggcg gcctcagtga gcgagcgagc gcgcagctgc
     4921 attaatgaat cggccaacgc gcggggagag gcggtttgcg tattgggcgc tcttccgctt
     4981 cctcgctcac tgactcgctg cgctcggtcg ttcggctgcg gcgagcggta tcagctcact
     5041 caaaggcggt aatacggtta tccacagaat caggggataa cgcaggaaag aacatgtgag
     5101 caaaaggcca gcaaaaggcc aggaaccgta aaaaggccgc gttgctggcg tttttccata
     5161 ggctccgccc ccctgacgag catcacaaaa atcgacgctc aagtcagagg tggcgaaacc
     5221 cgacaggact ataaagatac caggcgtttc cccctggaag ctccctcgtg cgctctcctg
     5281 ttccgaccct gccgcttacc ggatacctgt ccgcctttct cccttcggga agcgtggcgc
     5341 tttctcatag ctcacgctgt aggtatctca gttcggtgta ggtcgttcgc tccaagctgg
     5401 gctgtgtgca cgaacccccc gttcagcccg accgctgcgc cttatccggt aactatcgtc
     5461 ttgagtccaa cccggtaaga cacgacttat cgccactggc agcagccact ggtaacagga
     5521 ttagcagagc gaggtatgta ggcggtgcta cagagttctt gaagtggtgg cctaactacg
     5581 gctacactag aagaacagta tttggtatct gcgctctgct gaagccagtt accttcggaa
     5641 aaagagttgg tagctcttga tccggcaaac aaaccaccgc tggtagcggt ggtttttttg
     5701 tttgcaagca gcagattacg cgcagaaaaa aaggatctca agaagatcct ttgatctttt
     5761 ctacggggtc tgacgctcag tggaacgaaa actcacgtta agggattttg gtcatgagat
     5821 tatcaaaaag gatcttcacc tagatccttt taaattaaaa atgaagtttt aaatcaatct
     5881 aaagtatata tgagtaaact tggtctgaca gttaccaatg cttaatcagt gaggcaccta
     5941 tctcagcgat ctgtctattt cgttcatcca tagttgcctg actccccgtc gtgtagataa
     6001 ctacgatacg ggagggctta ccatctggcc ccagtgctgc aatgataccg cgagacccac
     6061 gctcaccggc tccagattta tcagcaataa accagccagc cggaagggcc gagcgcagaa
     6121 gtggtcctgc aactttatcc gcctccatcc agtctattaa ttgttgccgg gaagctagag
     6181 taagtagttc gccagttaat agtttgcgca acgttgttgc cattgctaca ggcatcgtgg
     6241 tgtcacgctc gtcgtttggt atggcttcat tcagctccgg ttcccaacga tcaaggcgag
     6301 ttacatgatc ccccatgttg tgcaaaaaag cggttagctc cttcggtcct ccgatcgttg
     6361 tcagaagtaa gttggccgca gtgttatcac tcatggttat ggcagcactg cataattctc
     6421 ttactgtcat gccatccgta agatgctttt ctgtgactgg tgagtactca accaagtcat
     6481 tctgagaata gtgtatgcgg cgaccgagtt gctcttgccc ggcgtcaata cgggataata
     6541 ccgcgccaca tagcagaact ttaaaagtgc tcatcattgg aaaacgttct tcggggcgaa
     6601 aactctcaag gatcttaccg ctgttgagat ccagttcgat gtaacccact cgtgcaccca
     6661 actgatcttc agcatctttt actttcacca gcgtttctgg gtgagcaaaa acaggaaggc
     6721 aaaatgccgc aaaaaaggga ataagggcga cacggaaatg ttgaatactc atactcttcc
     6781 tttttcaata ttattgaagc atttatcagg gttattgtct catgagcgga tacatatttg
     6841 aatgtattta gaaaaataaa caaatagggg ttccgcgcac atttccccga aaagtgccac
     6901 ctaaattgta agcgttaata ttttgttaaa attcgcgtta aatttttgtt aaatcagctc
     6961 attttttaac caataggccg aaatcggcaa aatcccttat aaatcaaaag aatagaccga
     7021 gatagggttg agtgttgttc cagtttggaa caagagtcca ctattaaaga acgtggactc
     7081 caacgtcaaa gggcgaaaaa ccgtctatca gggcgatggc ccactacgtg aaccatcacc
     7141 ctaatcaagt tttttggggt cgaggtgccg taaagcacta aatcggaacc ctaaagggag
     7201 cccccgattt agagcttgac ggggaaagcc ggcgaacgtg gcgagaaagg aagggaagaa
     7261 agcgaaagga gcgggcgcta gggcgctggc aagtgtagcg gtcacgctgc gcgtaaccac
     7321 cacacccgcc gcgcttaatg cgccgctaca gggcgcgtcc cattcgccat tcaggctg
//
