LOCUS       sequence-466527-        7382 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cation channelrhodopsin ChroME 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 7382)
  AUTHORS   Mardinly AR, Oldenburg IA, Pegard NC, Sridharan S, Lyall EH, Chesnov
            K, Brohawn SG, Waller L, Adesnik H
  TITLE     Precise multimodal optical control of neural ensemble activity.
  JOURNAL   Nat Neurosci. 2018 Apr 30. pii: 10.1038/s41593-018-0139-8. doi: 
            10.1038/s41593-018-0139-8.
  PUBMED    29713079
REFERENCE   2  (bases 1 to 7382)
  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 
            Neurosci. 2018 Apr 30. pii: 10.1038/s41593-018-0139-8. doi: 
            10.1038/s41593-018-0139-8."
COMMENT     Sequence Label: pAAV-CAG-DIO-ChroME-ST-P2A-H2B-mRuby3
FEATURES             Location/Qualifiers
     source          1..7382
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     rep_origin      1..589
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     490..509
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     repeat_region   826..955
                     /label=AAV2 ITR
     polyA_signal    1117..1238
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1154..1173)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1208..1227
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     1267..1283
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     primer_bind     1268..1284
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     misc_feature    1287..1875
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     CDS             1393..1404
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     1802..1822
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     protein_bind    complement(1900..1933)
                     /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(1984..2017)
                     /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             3017..3040
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     CDS             3251..3307
                     /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             3314..3691
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     CDS             3701..4411
                     /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"
     protein_bind    4423..4456
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     protein_bind    4507..4540
                     /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(4591..4610)
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     primer_bind     complement(4672..4694)
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     promoter        complement(4973..5251)
                     /label=chicken beta-actin promoter
     repeat_region   5565..5694
                     /label=AAV2 ITR
     rep_origin      5766..6221
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5853..5872)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     6063..6084
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        6247..6351
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6352..6420,6421..7212)
                     /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(6570..6589)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
ORIGIN
        1 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
       61 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
      121 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
      181 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
      241 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
      301 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
      361 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
      421 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
      481 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
      541 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
      601 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
      661 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
      721 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
      781 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctgcg cgctcgctcg
      841 ctcactgagg ccgcccgggc aaagcccggg cgtcgggcga cctttggtcg cccggcctca
      901 gtgagcgagc gagcgcgcag agagggagtg gccaactcca tcactagggg ttccttgtag
      961 ttaatgatta acccgccatg ctacttatct acgtagccat gctctaggaa gatccttatc
     1021 gattttacca catttgtaga ggttttactt gctttaaaaa acctcccaca tctccccctg
     1081 aacctgaaac ataaaatgaa tgcaattgtt gttgttaact tgtttattgc agcttataat
     1141 ggttacaaat aaagcaatag catcacaaat ttcacaaata aagcattttt ttcactgcat
     1201 tctagttgtg gtttgtccaa actcatcaat gtatcttatc atgtctgctc gaagcggccg
     1261 cccgggtcga ggtcgacggt atcgatgcgg ggaggcggcc caaagggaga tccgactcgt
     1321 ctgagggcga aggcgaagac gcggaagagg ccgcagagcc ggcagcaggc cgcgggaagg
     1381 aaggtccgct ggattgaggg ccgaagggac gtagcagaag gacgtcccgc gcagaatcca
     1441 ggtggcaaca caggcgagca gccaaggaaa ggacgatgat ttccccgaca acaccacgga
     1501 attgtcagtg cccaacagcc gagcccctgt ccagcagcgg gcaaggcagg cggcgatgag
     1561 ttccgccgtg gcaataggga gggggaaagc gaaagtcccg gaaaggagct gacaggtggt
     1621 ggcaatgccc caaccagtgg gggttgcgtc agcaaacaca gtgcacacca cgccacgttg
     1681 cctgacaacg ggccacaact cctcataaag agacagcaac caggatttat acaaggagga
     1741 gaaaatgaaa gccatacggg aagcaatagc atgatacaaa ggcattaaag cagcgtatcc
     1801 acatagcgta aaaggagcaa catagttaag aataccagtc aatctttcac aaattttgta
     1861 atccagaggt tgattatcga taagcttgat atcgaattca taacttcgta tagcatacat
     1921 tatacgaagt tattctttgc accattctaa agaataacag tgataatttc tgggttaagg
     1981 caaataactt cgtataggat actttatacg aagttatgct agcagtatgg aaacagccgc
     2041 cacaatgacc cacgccttta tctcagccgt gcctagcgcc gaagccacaa ttagaggcct
     2101 gctgagcgcc gcagcagtgg tgacaccagc agcagacgct cacggagaaa cctctaacgc
     2161 cacaacagcc ggagccgatc acggttgctt cccccacatc aaccacggaa ccgagctgca
     2221 gcacaagatc gcagtgggac tccagtggtt caccgtgatc gtggctatcg tgcagctcat
     2281 cttctacggt tggcacagct tcaaggccac aaccggctgg gaggaggtct acgtctgcgt
     2341 gatcgagctc gtcaagtgct tcatcgagct gttccacgag gtcgacagcc cagccacagt
     2401 gtaccagacc aacggaggag ccgtgatttg gctgcggtac agcgagtggc tcctgacttg
     2461 ccccgtgatc ctgatccacc tgagcaacct gaccggactg cacgaagagt acagcaagcg
     2521 gaccatgacc atcctggtga ccgacatcgg caacatcgtg tgggggatca cagccgcctt
     2581 tacaaagggc cccctgaaga tcctgttctt catgatcggc ctgttctacg gcgtgacttg
     2641 cttcttccag atcgccaagg tgtatatcga gagctaccac accctgccca aaggcgtctg
     2701 ccggaagatt tgcaagatca tggcctacgt cttcttctgc tcttggctga tgttccccgt
     2761 gatgttcatc gccggacacg agggactggg cctgatcaca ccttacacca gcggaatcgg
     2821 ccacctgatc ctggatctga tcagcaagaa cacttggggc ttcctgggcc accacctgag
     2881 agtgaagatc cacgagcaca tcctgatcca cggcgacatc cggaagacaa ccaccatcaa
     2941 cgtggccggc gagaacatgg agatcgagac cttcgtcgac gaggaggagg agggaggagt
     3001 ggcggccgca ccggtagatt acaaagacga tgacgataag cagtcccagc ccatcctcaa
     3061 caccaaggag atggccccgc agagcaagcc tccagaggag ctggagatga gcagcatgcc
     3121 cagccccgtg gcccctctgc ccgcacgcac ggagggcgtc atcgacatgc ggagcatgtc
     3181 cagcattgac agcttcatca gctgtgccac ggacttccct gaagccacca gattcgcggc
     3241 aggaagcgga gctactaact tcagcctgct gaagcaggct ggagacgtgg aggagaaccc
     3301 tggacctact agtatgccag agccagcgaa gtctgctccc gccccgaaaa agggctccaa
     3361 gaaggcggtg actaaggcgc agaagaaagg cggcaagaag cgcaagcgca gccgcaagga
     3421 gagctattcc atctatgtgt acaaggttct gaagcaggtc caccctgaca ccggcatttc
     3481 gtccaaggcc atgggcatca tgaattcgtt tgtgaacgac attttcgagc gcatcgcagg
     3541 tgaggcttcc cgcctggcgc attacaacaa gcgctcgacc atcacctcca gggagatcca
     3601 gacggccgtg cgcctgctgc tgcctgggga gttggccaag cacgccgtgt ccgagggtac
     3661 taaggccatc accaagtaca ccagcgctaa ggcactcgag atggtgtcta agggcgaaga
     3721 gctgatcaag gaaaatatgc gtatgaaggt ggtcatggaa ggttcggtca acggccacca
     3781 attcaaatgc acaggtgaag gagaaggcag accgtacgag ggagtgcaaa ccatgaggat
     3841 caaagtcatc gagggaggac ccctgccatt tgcctttgac attcttgcca cgtcgttcat
     3901 gtatggcagc cgtaccttta tcaagtaccc ggccgacatc cctgatttct ttaaacagtc
     3961 ctttcctgag ggttttactt gggaaagagt tacgagatac gaagatggtg gagtcgtcac
     4021 cgtcacgcag gacaccagcc ttgaggatgg cgagctcgtc tacaacgtca aggtcagagg
     4081 ggtaaacttt ccctccaatg gtcccgtgat gcagaagaag accaagggtt gggagcctaa
     4141 tacagagatg atgtatccag cagatggtgg tctgagagga tacactgaca tcgcactgaa
     4201 agttgatggt ggtggccatc tgcactgcaa cttcgtgaca acttacaggt caaaaaagac
     4261 cgtcgggaac atcaagatgc ccggtgtcca tgccgttgat caccgcctgg aaaggatcga
     4321 ggagagtgac aatgaaacct acgtagtgca aagagaagtg gcagttgcca aatacagcaa
     4381 ccttggtggt ggcatggacg agctgtacaa gtgaggcgcg ccataacttc gtataatgta
     4441 tgctatacga agttattaag aggtttcata ttgctaatag cagctacaat ccagctacca
     4501 ttctgcataa cttcgtataa agtatcctat acgaagttat tccggagtcg actctagagg
     4561 atccaattct ttgccaaaat gatgagacag cacaataacc agcacgttgc ccaggagctg
     4621 taggaaaaag aagaaggcat gaacatggtt agcagaggct ctagagccgc cggtcacacg
     4681 ccagaagccg aaccccgccc tgccccgtcc cccccgaagg cagccgtccc cccgcggaca
     4741 gccccgagcc gctccccccg cacaaagggc cctcccggag cccctcaagg ctttcacgca
     4801 gccacagaaa agaaacaagc cgtcattaaa ccaagcgcta attacagccc ggaggagaag
     4861 ggccgtcccg cccgctcacc tgtgggagta acgcggtcag tcagagccgg ggcgggcggc
     4921 gcgaggcggc ggcggagcgg ggcacggggc gaaggcagcg cgcagcgact cccgcccgcc
     4981 gcgcgcttcg ctttttatag ggccgccgcc gccgccgcct cgccataaaa ggaaactttc
     5041 ggagcgcgcc gctctgattg gctgccgccg cacctctccg cctcgccccg ccccgcccct
     5101 cgccccgccc cgccccgcct ggcgcgcgcc cccccccccc cccccgcccc catcgctgca
     5161 caaaataatt aaaaaataaa taaatacaaa attgggggtg gggagggggg ggagatgggg
     5221 agagtgaagc agaacgtggg gctcacctcg accatggtaa tagcgatgac taatacgtag
     5281 atgtactgcc aagtaggaaa gtcccataag gtcatgtact gggcataatg ccaggcgggc
     5341 catttaccgt cattgacgtc aatagggggc gtacttggca tatgatacac ttgatgtact
     5401 gccaagtggg cagtttaccg taaatactcc acccattgac gtcaatggaa agtccctatt
     5461 ggcgttacta tgggaacata cgtcattatt gacgtcaatg gtacagatct tcctagagca
     5521 tggctacgta gataagtagc atggcgggtt aatcattaac tacaaggaac ccctagtgat
     5581 ggagttggcc actccctctc tgcgcgctcg ctcgctcact gaggccgggc gaccaaaggt
     5641 cgcccgacgc ccgggctttg cccgggcggc ctcagtgagc gagcgagcgc gcagctggcg
     5701 taatagcgaa gaggcccgca ccgatcgccc ttcccaacag ttgcgcagcc tgaatggcga
     5761 atgggacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     5821 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
     5881 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     5941 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag
     6001 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     6061 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga
     6121 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
     6181 atttaacgcg aattttaaca aaatattaac gcttacaatt taggtggcac ttttcgggga
     6241 aatgtgcgcg gaacccctat ttgtttattt ttctaaatac attcaaatat gtatccgctc
     6301 atgagacaat aaccctgata aatgcttcaa taatattgaa aaaggaagag tatgagtatt
     6361 caacatttcc gtgtcgccct tattcccttt tttgcggcat tttgccttcc tgtttttgct
     6421 cacccagaaa cgctggtgaa agtaaaagat gctgaagatc agttgggtgc acgagtgggt
     6481 tacatcgaac tggatctcaa cagcggtaag atccttgaga gttttcgccc cgaagaacgt
     6541 tttccaatga tgagcacttt taaagttctg ctatgtggcg cggtattatc ccgtattgac
     6601 gccgggcaag agcaactcgg tcgccgcata cactattctc agaatgactt ggttgagtac
     6661 tcaccagtca cagaaaagca tcttacggat ggcatgacag taagagaatt atgcagtgct
     6721 gccataacca tgagtgataa cactgcggcc aacttacttc tgacaacgat cggaggaccg
     6781 aaggagctaa ccgctttttt gcacaacatg ggggatcatg taactcgcct tgatcgttgg
     6841 gaaccggagc tgaatgaagc cataccaaac gacgagcgtg acaccacgat gcctgtagca
     6901 atggcaacaa cgttgcgcaa actattaact ggcgaactac ttactctagc ttcccggcaa
     6961 caattaatag actggatgga ggcggataaa gttgcaggac cacttctgcg ctcggccctt
     7021 ccggctggct ggtttattgc tgataaatct ggagccggtg agcgtgggtc tcgcggtatc
     7081 attgcagcac tggggccaga tggtaagccc tcccgtatcg tagttatcta cacgacgggg
     7141 agtcaggcaa ctatggatga acgaaataga cagatcgctg agataggtgc ctcactgatt
     7201 aagcattggt aactgtcaga ccaagtttac tcatatatac tttagattga tttaaaactt
     7261 catttttaat ttaaaaggat ctaggtgaag atcctttttg ataatctcat gaccaaaatc
     7321 ccttaacgtg agttttcgtt ccactgagcg tcagaccccg tagaaaagat caaaggatct
     7381 tc
//
