LOCUS       sequence-338751-        7696 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Cation channelrhodopsin ChroME2f 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 7696)
  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 7696)
  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-ChroME2f-ST-P2A-H2B-mRuby3
FEATURES             Location/Qualifiers
     source          1..7696
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     repeat_region   383..512
                     /label=AAV2 ITR
     primer_bind     1382..1404
                     /label=pCAGGS-5
                     /note="Chimeric intron in CAG promoter, forward primer"
     primer_bind     1466..1485
                     /label=pCAG-F
                     /note="Rabbit beta-globin intron, for pCAG plasmids, 
                     forward primer"
     protein_bind    complement(1536..1569)
                     /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(1620..1653)
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     CDS             complement(1665..2375)
                     /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(2385..2762)
                     /codon_start=1
                     /gene="HIST1H2BJ"
                     /product="human histone H2B"
                     /label=H2B
                     /translation="MPEPAKSAPAPKKGSKKAVTKAQKKGGKKRKRSRKESYSIYVYKV
                     LKQVHPDTGISSKAMGIMNSFVNDIFERIAGEASRLAHYNKRSTITSREIQTAVRLLLP
                     GELAKHAVSEGTKAITKYTSAK"
     CDS             complement(2769..2825)
                     /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(3036..3059)
                     /codon_start=1
                     /product="FLAG(R) epitope tag, followed by an enterokinase 
                     cleavage site"
                     /label=FLAG
                     /translation="DYKDDDDK"
     protein_bind    4059..4092
                     /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    4143..4176
                     /label=loxP
                     /bound_moiety="Cre recombinase"
                     /note="Cre-mediated recombination occurs in the 8-bp core 
                     sequence (ATGTATGC) (Shaw et al., 2021)."
     misc_feature    4201..4789
                     /label=WPRE
                     /note="woodchuck hepatitis virus posttranscriptional 
                     regulatory element"
     primer_bind     complement(4254..4274)
                     /label=WPRE-R
                     /note="WPRE, reverse primer"
     CDS             complement(4672..4683)
                     /codon_start=1
                     /product="Factor Xa recognition and cleavage site"
                     /label=Factor Xa site
                     /translation="IEGR"
     primer_bind     complement(4792..4808)
                     /label=KS primer
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(4793..4809)
                     /label=pBluescriptKS
                     /note="For pBluescript vector"
     polyA_signal    4838..4959
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4849..4868)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     4903..4922
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     repeat_region   5121..5250
                     /label=AAV2 ITR
     rep_origin      5431..5886
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(5518..5537)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     5728..5749
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     promoter        5912..6016
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(6017..6085,6086..6877)
                     /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(6235..6254)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      7048..7636
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     7537..7556
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
ORIGIN
        1 ttctttcctg cgttatcccc tgattctgtg gataaccgta ttaccgcctt tgagtgagct
       61 gataccgctc gccgcagccg aacgaccgag cgcagcgagt cagtgagcga ggaagcggaa
      121 gagcgcccaa tacgcaaacc gcctctcccc gcgcgttggc cgattcatta atgcagctgc
      181 gcgctcgctc gctcactgag gccgcccggg caaagcccgg gcgtcgggcg acctttggtc
      241 gcccggcctc agtgagcgag cgagcgcgca gagagggagt ggccmrctym atsmsymgct
      301 acagggcgcg tcccattcgc cattcaggct gcgcaactgt tgggaagggc gatcggtgcg
      361 ggcctcttcg ctattacgcc agctgcgcgc tcgctcgctc actgaggccg cccgggcaaa
      421 gcccgggcgt cgggcgacct ttggtcgccc ggcctcagtg agcgagcgag cgcgcagaga
      481 gggagtggcc aactccatca ctaggggttc cttgtagtta atgattaacc cgccatgcta
      541 cttatctacg tagccatgct ctaggaagat ctgtaccatt gacgtcaata atgacgtatg
      601 ttcccatagt aacgccaata gggactttcc attgacgtca atgggtggag tatttacggt
      661 aaactgccca cttggcagta catcaagtgt atcatatgcc aagtacgccc cctattgacg
      721 tcaatgacgg taaatggccc gcctggcatt atgcccagta catgacctta tgggactttc
      781 ctacttggca gtacatctac gtattagtca tcgctattac catggtcgag gtgagcccca
      841 cgttctgctt cactctcccc atctcccccc cctccccacc cccaattttg tatttattta
      901 ttttttaatt attttgtgca gcgatggggg cggggggggg gggggggsgc gcgccggggg
      961 gggggggggg gggsgggggg gggggsgggg sgrggcggag aggkgcggcg gcagccaatc
     1021 agagcggcgc gctccgaaag tttcctttta tggcgaggcg gcggcggcgg cggccctata
     1081 aaaagcgaag cgcgcggcgg gcgggagtcg ctgcgcgctg ccttcgcccc gtgccccgct
     1141 ccgccgccgc ctcgcgccgc ccgccccggc tctgactgac cgcgttactc ccacaggtga
     1201 gcgggcggga cggcccttct cctccgggct gtaattagcg cttggtttaa tgacggcttg
     1261 tttcttttct gtggctgcgt gaaagccttg aggggctccg ggagggccct ttgtgcgggg
     1321 ggagcggctc ggggctgtcc gcggggggac ggctgccttc gggggggacg gggcagggcg
     1381 gggttcggct tctggcgtgt gaccggcggc tctagagcct ctgctaacca tgttcatgcc
     1441 ttcttctttt tcctacagct cctgggcaac gtgctggtta ttgtgctgtc tcatcatttt
     1501 ggcaaagaat tggatcctct agagtcgact ccggaataac ttcgtatagg atactttata
     1561 cgaagttatg cagaatggta gctggattgt agctgctatt agcaatatga aacctcttaa
     1621 taacttcgta tagcatacat tatacgaagt tatggcgcgc ctcacttgta cagctcgtcc
     1681 atgccaccac caaggttgct gtatttggca actgccactt ctctttgcac tacgtaggtt
     1741 tcattgtcac tctcctcgat cctttccagg cggtgatcaa cggcatggac accgggcatc
     1801 ttgatgttcc cgacggtctt ttttgacctg taagttgtca cgaagttgca gtgcagatgg
     1861 ccaccaccat caactttcag tgcgatgtca gtgtatcctc tcagaccacc atctgctgga
     1921 tacatcatct ctgtattagg ctcccaaccc ttggtcttct tctgcatcac gggaccattg
     1981 gagggaaagt ttacccctct gaccttgacg ttgtagacga gctcgccatc ctcaaggctg
     2041 gtgtcctgcg tgacggtgac gactccacca tcttcgtatc tcgtaactct ttcccaagta
     2101 aaaccctcag gaaaggactg tttaaagaaa tcagggatgt cggccgggta cttgataaag
     2161 gtacggctgc catacatgaa cgacgtggca agaatgtcaa aggcaaatgg caggggtcct
     2221 ccctcgatga ctttgatcct catggtttgc actccctcgt acggtctgcc ttctccttca
     2281 cctgtgcatt tgaattggtg gccgttgacc gaaccttcca tgaccacctt catacgcata
     2341 ttttccttga tcagctcttc gcccttagac accatctcga gtgccttagc gctggtgtac
     2401 ttggtgatgg ccttagtacc ctcggacacg gcgtgcttgg ccaactcccc aggcagcagc
     2461 aggcgcacgg ccgtctggat ctccctggag gtgatggtcg agcgcttgtt gtaatgcgcc
     2521 aggcgggaag cctcacctgc gatgcgctcg aaaatgtcgt tcacaaacga attcatgatg
     2581 cccatggcct tggacgaaat gccggtgtca gggtggacct gcttcagaac cttgtacaca
     2641 tagatggaat agctctcctt gcggctgcgc ttgcgcttct tgccgccttt cttctgcgcc
     2701 ttagtcaccg ccttcttgga gccctttttc ggggcgggag cagacttcgc tggctctggc
     2761 atactagtag gtccagggtt ctcctccacg tctccagcct gcttcagcag gctgaagtta
     2821 gtagctccgc ttcctgccgc gaatctggtg gcttcaggga agtccgtggc acagctgatg
     2881 aagctgtcaa tgctggacat gctccgcatg tcgatgacgc cctccgtgcg tgcgggcaga
     2941 ggggccacgg ggctgggcat gctgctcatc tccagctcct ctggaggctt gctctgcggg
     3001 gccatctcct tggtgttgag gatgggctgg gactgcttat cgtcatcgtc tttgtaatct
     3061 accggtgcgg ccgccactcc tccctcctcc tcctcgtcga cgaaggtctc gatctccatg
     3121 ttctcgccgg ccacgttgat ggtggttgtc ttccggatgt cgccgtggat caggatgtgc
     3181 tcgtggatct tcactctcag gtggtggccc aggaagcccc aagtgttctt ggcgatcaga
     3241 tccaggatca ggtggccgat tccgctggtg taaggtgtga tcaggcccag tccctcgtgt
     3301 ccggcgatga acatcacggg gaacatcagc caagagcaga agaagacgta ggccatgatc
     3361 ttgcaaatct tccggcagac gcctttgggc agggtgtggt agctctcgat atacaccttg
     3421 gcgatctgga agaagcaagt cacgccgtag aacaggccga tcatgaagaa caggatcttc
     3481 agggggccct ttgtaaaggc ggctgtgatc ccccacacga tgttgccgat gtcggtcacc
     3541 aggatggtca tggtccgctt gctgtactct tcgtgcagtc cggtcaggtt gctcaggtgg
     3601 atcaggatca cggggcaagt caggagccac tcgctgtacc gcagccaagc cacggctcct
     3661 ccgttggtct ggtacactgt ggctgggctg tcgacatcgt ggaacagctc gatgaagcac
     3721 ttgacgagct cgatcacgca gacgtagacc tcctcccagc cggttgtggc cttgaagctg
     3781 tgccaaccgt agaagatgag ctgcacgata gccacgatca cggtgaacca ctggagtccc
     3841 actgcgatct tgtgctgcag ctcggttccg tggttgatgt gggggaagca accgtgatcg
     3901 gctccggctg ttgtggcgtt agaggtttct ccgtgagcgt ctgctgctgg tgtcaccact
     3961 gctgcggcgc tcagcaggcc tctaattgtg gcttcggcgc taggcacggc tgagataaag
     4021 gcgtgggtca ttgtggcggc tgtttccata ctgctagcat aacttcgtat aaagtatcct
     4081 atacgaagtt atttgcctta acccagaaat tatcactgtt attctttaga atggtgcaaa
     4141 gaataacttc gtataatgta tgctatacga agttatgaat tcgatatcaa gcttatcgat
     4201 aatcaacctc tggattacaa aatttgtgaa agattgactg gtattcttaa ctatgttgct
     4261 ccttttacgc tatgtggata cgctgcttta atgcctttgt atcatgctat tgcttcccgt
     4321 atggctttca ttttctcctc cttgtataaa tcctggttgc tgtctcttta tgaggagttg
     4381 tggcccgttg tcaggcaacg tggcgtggtg tgcactgtgt ttgctgacgc aacccccact
     4441 ggttggggca ttgccaccac ctgtcagctc ctttccggga ctttcgcttt ccccctccct
     4501 attgccacgg cggaactcat cgccgcctgc cttgcccgct gctggacagg ggctcggctg
     4561 ttgggcactg acaattccgt ggtgttgtcg gggaaatcat cgtcctttcc ttggctgctc
     4621 gcctgtgttg ccacctggat tctgcgcggg acgtccttct gctacgtccc ttcggccctc
     4681 aatccagcgg accttccttc ccgcggcctg ctgccggctc tgcggcctct tccgcgtctt
     4741 cgccttcgcc ctcagacgag tcggatctcc ctttgggccg cctccccgca tcgataccgt
     4801 cgacctcgac ccgggcggcc gcttcgagca gacatgataa gatacattga tgagtttgga
     4861 caaaccacaa ctagaatgca gtgaaaaaaa tgctttattt gtgaaatttg tgatgctatt
     4921 gctttatttg taaccattat aagctgcaat aaacaagtta acaacaacaa ttgcattcat
     4981 tttatgtttc aggttcaggg ggagatgtgg gaggtttttt aaagcaagta aaacctctac
     5041 aaatgtggta aaatcgataa ggatcttcct agagcatggc tacgtagata agtagcatgg
     5101 cgggttaatc attaactaca aggaacccct agtgatggag ttggccactc cctctctgcg
     5161 cgctcgctcg ctcactgagg ccgggcgacc aaaggtcgcc cgacgcccgg gctttgcccg
     5221 ggcggcctca gtgagcgagc gagcgcgcag ctgcattaat gaatcggccm ackcgcksgs
     5281 wsastgaggc cgggcgacca aaggtcgccc gacgcccggg ctttgcccgg gcggcctcag
     5341 tgagcgagcg agcgcgcagc tggcgtaata gcgaagaggc ccgcaccgat cgcccttccc
     5401 aacagttgcg cagcctgaat ggcgaatggg acgcgccctg tagcggcgca ttaagcgcgg
     5461 cgggtgtggt ggttacgcgc agcgtgaccg ctacacttgc cagcgcccta gcgcccgctc
     5521 ctttcgcttt cttcccttcc tttctcgcca cgttcgccgg ctttccccgt caagctctaa
     5581 atcgggggct ccctttaggg ttccgattta gtgctttacg gcacctcgac cccaaaaaac
     5641 ttgattaggg tgatggttca cgtagtgggc catcgccctg atagacggtt tttcgccctt
     5701 tgacgttgga gtccacgttc tttaatagtg gactcttgtt ccaaactgga acaacactca
     5761 accctatctc ggtctattct tttgatttat aagggatttt gccgatttcg gcctattggt
     5821 taaaaaatga gctgatttaa caaaaattta acgcgaattt taacaaaata ttaacgctta
     5881 caatttaggt ggcacttttc ggggaaatgt gcgcggaacc cctatttgtt tatttttcta
     5941 aatacattca aatatgtatc cgctcatgag acaataaccc tgataaatgc ttcaataata
     6001 ttgaaaaagg aagagtatga gtattcaaca tttccgtgtc gcccttattc ccttttttgc
     6061 ggcattttgc cttcctgttt ttgctcaccc agaaacgctg gtgaaagtaa aagatgctga
     6121 agatcagttg ggtgcacgag tgggttacat cgaactggat ctcaacagcg gtaagatcct
     6181 tgagagtttt cgccccgaag aacgttttcc aatgatgagc acttttaaag ttctgctatg
     6241 tggcgcggta ttatcccgta ttgacgccgg gcaagagcaa ctcggtcgcc gcatacacta
     6301 ttctcagaat gacttggttg agtactcacc agtcacagaa aagcatctta cggatggcat
     6361 gacagtaaga gaattatgca gtgctgccat aaccatgagt gataacactg cggccaactt
     6421 acttctgaca acgatcggag gaccgaagga gctaaccgct tttttgcaca acatggggga
     6481 tcatgtaact cgccttgatc gttgggaacc ggagctgaat gaagccatac caaacgacga
     6541 gcgtgacacc acgatgcctg tagcaatggc aacaacgttg cgcaaactat taactggcga
     6601 actacttact ctagcttccc ggcaacaatt aatagactgg atggaggcgg ataaagttgc
     6661 aggaccactt ctgcgctcgg cccttccggc tggctggttt attgctgata aatctggagc
     6721 cggtgagcgt gggtctcgcg gtatcattgc agcactgggg ccagatggta agccctcccg
     6781 tatcgtagtt atctacacga cggggagtca ggcaactatg gatgaacgaa atagacagat
     6841 cgctgagata ggtgcctcac tgattaagca ttggtaactg tcagaccaag tttactcata
     6901 tatactttag attgatttaa aacttcattt ttaatttaaa aggatctagg tgaagatcct
     6961 ttttgataat ctcatgacca aaatccctta acgtgagttt tcgttccact gagcgtcaga
     7021 ccccgtagaa aagatcaaag gatcttcttg agatcctttt tttctgcgcg taatctgctg
     7081 cttgcaaaca aaaaaaccac cgctaccagc ggtggtttgt ttgccggatc aagagctacc
     7141 aactcttttt ccgaaggtaa ctggcttcag cagagcgcag ataccaaata ctgttcttct
     7201 agtgtagccg tagttaggcc accacttcaa gaactctgta gcaccgccta catacctcgc
     7261 tctgctaatc ctgttaccag tggctgctgc cagtggcgat aagtcgtgtc ttaccgggtt
     7321 ggactcaaga cgatagttac cggataaggc gcagcggtcg ggctgaacgg ggggttcgtg
     7381 cacacagccc agcttggagc gaacgaccta caccgaactg agatacctac agcgtgagct
     7441 atgagaaagc gccacgcttc ccgaagggag aaaggcggac aggtatccgg taagcggcag
     7501 ggtcggaaca ggagagcgca cgagggagct tccaggggga aacgcctggt atctttatag
     7561 tcctgtcggg tttcgccacc tctgacttga gcgtcgattt ttgtgatgct cgtcaggggg
     7621 gcggagccta tggaaaaacg ccagcaacgc ggccttttta cggttcctgg ccttttgctg
     7681 gccttttgct cacatg
//
