LOCUS       sequence-181022-        7190 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Mammalian expression of humanized ChR2 with H134R mutation fused to 
            mCherry for optogenetic activation.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7190)
  AUTHORS   Zhang F, Wang LP, Brauner M, Liewald JF, Kay K, Watzke N, Wood PG, 
            Bamberg E, Nagel G, Gottschalk A, Deisseroth K
  TITLE     Multimodal fast optical interrogation of neural circuitry.
  JOURNAL   Nature. 2007 Apr 5. 446(7136):633-9.
  PUBMED    17410168
REFERENCE   2  (bases 1 to 7190)
  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: "Nature. 
            2007 Apr 5. 446(7136):633-9."
COMMENT     Sequence Label: pcDNA3.1/hChR2(H134R)-mCherry
FEATURES             Location/Qualifiers
     source          1..7190
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(46..65)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        237..616
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        617..820
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     771..791
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     865..884
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        865..883
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     regulatory      959..968
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             965..1891
                     /codon_start=1
                     /product="H134R mutant of an N-terminal fragment of 
                     channelrhodopsin-2 from the alga Chlamydomonas reinhardtii 
                     (Nagel et al., 2005)"
                     /label=ChR2(H134R)
                     /note="blue light-activated cation channel"
                     /translation="MDYGGALSAVGRELLFVTNPVVVNGSVLVPEDQCYCAGWIESRGT
                     NGAQTASNVLQWLAAGFSILLLMFYAYQTWKSTCGWEEIYVCAIEMVKVILEFFFEFKN
                     PSMLYLATGHRVQWLRYAEWLLTCPVILIRLSNLTGLSNDYSRRTMGLLVSDIGTIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     CDS             1904..2611
                     /codon_start=1
                     /product="monomeric derivative of DsRed fluorescent protein
                     (Shaner et al., 2004)"
                     /label=mCherry
                     /note="mammalian codon-optimized"
                     /translation="VSKGEEDNMAIIKEFMRFKVHMEGSVNGHEFEIEGEGEGRPYEGT
                     QTAKLKVTKGGPLPFAWDILSPQFMYGSKAYVKHPADIPDYLKLSFPEGFKWERVMNFE
                     DGGVVTVTQDSSLQDGEFIYKVKLRGTNFPSDGPVMQKKTMGWEASSERMYPEDGALKG
                     EIKQRLKLKDGGHYDAEVKTTYKAKKPVQLPGAYNVNIKLDITSHNEDYTIVEQYERAE
                     GRHSTGGMDELYK"
     primer_bind     complement(2046..2064)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2314..2333
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     CDS             2690..2731
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             2741..2758
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(2781..2798)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    2787..3011
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      3057..3485
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3144..3163)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     3354..3375
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(3494..3514)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        3499..3828
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      3679..3814
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     3741..3760
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             3895..4689
                     /codon_start=1
                     /gene="aph(3')-II (or nptII)"
                     /product="aminoglycoside phosphotransferase from Tn5"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKARMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(3949..3968)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4559..4578
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    4865..4986
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(4902..4921)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     4956..4975
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(5035..5051)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(5035..5051)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(5048..5070)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    5059..5075
                     /label=lac operator
                     /bound_moiety="lac repressor encoded by lacI"
                     /note="The lac repressor binds to the lac operator to 
                     inhibit transcription in E. coli. This inhibition can be 
                     relieved by adding lactose or 
                     isopropyl-beta-D-thiogalactopyranoside (IPTG)."
     promoter        complement(join(5083..5089,5090..5107,5108..5113))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5128..5149
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(5266..5283)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(5437..6025)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(5517..5536)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(6196..6987,6988..7056))
                     /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     6819..6838
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(7057..7161)
                     /gene="bla"
                     /label=AmpR promoter
ORIGIN
        1 tcgacggatc gggagatctc ccgatcccct atggtgcact ctcagtacaa tctgctctga
       61 tgccgcatag ttaagccagt atctgctccc tgcttgtgtg ttggaggtcg ctgagtagtg
      121 cgcgagcaaa atttaagcta caacaaggca aggcttgacc gacaattgca tgaagaatct
      181 gcttagggtt aggcgttttg cgctgcttcg cgatgtacgg gccagatata cgcgttgaca
      241 ttgattattg actagttatt aatagtaatc aattacgggg tcattagttc atagcccata
      301 tatggagttc cgcgttacat aacttacggt aaatggcccg cctggctgac cgcccaacga
      361 cccccgccca ttgacgtcaa taatgacgta tgttcccata gtaacgccaa tagggacttt
      421 ccattgacgt caatgggtgg agtatttacg gtaaactgcc cacttggcag tacatcaagt
      481 gtatcatatg ccaagtacgc cccctattga cgtcaatgac ggtaaatggc ccgcctggca
      541 ttatgcccag tacatgacct tatgggactt tcctacttgg cagtacatct acgtattagt
      601 catcgctatt accatggtga tgcggttttg gcagtacatc aatgggcgtg gatagcggtt
      661 tgactcacgg ggatttccaa gtctccaccc cattgacgtc aatgggagtt tgttttggca
      721 ccaaaatcaa cgggactttc caaaatgtcg taacaactcc gccccattga cgcaaatggg
      781 cggtaggcgt gtacggtggg aggtctatat aagcagagct ctctggctaa ctagagaacc
      841 cactgcttac tggcttatcg aaattaatac gactcactat agggagaccc aagctggcta
      901 gttaagcttg gtaccgagct cggatccact agtccagtgt ggtggaattg cccttgctgc
      961 caccatggac tatggcggcg ctttgtctgc cgtcggacgc gaacttttgt tcgttactaa
     1021 tcctgtggtg gtgaacgggt ccgtcctggt ccctgaggat caatgttact gtgccggatg
     1081 gattgaatct cgcggcacga acggcgctca gaccgcgtca aatgtcctgc agtggcttgc
     1141 agcaggattc agcattttgc tgctgatgtt ctatgcctac caaacctgga aatctacatg
     1201 cggctgggag gagatctatg tgtgcgccat tgaaatggtt aaggtgattc tcgagttctt
     1261 ttttgagttt aagaatccct ctatgctcta ccttgccaca ggacaccggg tgcagtggct
     1321 gcgctatgca gagtggctgc tcacttgtcc tgtcatcctt atccgcctga gcaacctcac
     1381 cggcctgagc aacgactaca gcaggagaac catgggactc cttgtctcag acatcgggac
     1441 tatcgtgtgg ggggctacca gcgccatggc aaccggctat gttaaagtca tcttcttttg
     1501 tcttggattg tgctatggcg cgaacacatt ttttcacgcc gccaaagcat atatcgaggg
     1561 ttatcatact gtgccaaagg gtcggtgccg ccaggtcgtg accggcatgg catggctgtt
     1621 tttcgtgagc tggggtatgt tcccaattct cttcattttg gggcccgaag gttttggcgt
     1681 cctgagcgtc tatggctcca ccgtaggtca cacgattatt gatctgatga gtaaaaattg
     1741 ttgggggttg ttgggacact acctgcgcgt cctgatccac gagcacatat tgattcacgg
     1801 agatatccgc aaaaccacca aactgaacat cggcggaacg gagatcgagg tcgagactct
     1861 cgtcgaagac gaagccgagg ccggagccgt gccagcggcc gccgtgagca agggcgagga
     1921 ggataacatg gccatcatca aggagttcat gcgcttcaag gtgcacatgg agggctccgt
     1981 gaacggccac gagttcgaga tcgagggcga gggcgagggc cgcccctacg agggcaccca
     2041 gaccgccaag ctgaaggtga ccaagggtgg ccccctgccc ttcgcctggg acatcctgtc
     2101 ccctcagttc atgtacggct ccaaggccta cgtgaagcac cccgccgaca tccccgacta
     2161 cttgaagctg tccttccccg agggcttcaa gtgggagcgc gtgatgaact tcgaggacgg
     2221 cggcgtggtg accgtgaccc aggactcctc cctgcaggac ggcgagttca tctacaaggt
     2281 gaagctgcgc ggcaccaact tcccctccga cggccccgta atgcagaaga agaccatggg
     2341 ctgggaggcc tcctccgagc ggatgtaccc cgaggacggc gccctgaagg gcgagatcaa
     2401 gcagaggctg aagctgaagg acggcggcca ctacgacgct gaggtcaaga ccacctacaa
     2461 ggccaagaag cccgtgcagc tgcccggcgc ctacaacgtc aacatcaagt tggacatcac
     2521 ctcccacaac gaggactaca ccatcgtgga acagtacgaa cgcgccgagg gccgccactc
     2581 caccggcggc atggacgagc tgtacaagta atgagtcgac gccaagggca attctgcaga
     2641 tatccagcac agtggcggcc gctcgagtct agagggcccg cggttcgaag gtaagcctat
     2701 ccctaaccct ctcctcggtc tcgattctac gcgtaccggt catcatcacc atcaccattg
     2761 agtttaaacc cgctgatcag cctcgactgt gccttctagt tgccagccat ctgttgtttg
     2821 cccctccccc gtgccttcct tgaccctgga aggtgccact cccactgtcc tttcctaata
     2881 aaatgaggaa attgcatcgc attgtctgag taggtgtcat tctattctgg ggggtggggt
     2941 ggggcaggac agcaaggggg aggattggga agacaatagc aggcatgctg gggatgcggt
     3001 gggctctatg gcttctgagg cggaaagaac cagctggggc tctagggggt atccccacgc
     3061 gccctgtagc ggcgcattaa gcgcggcggg tgtggtggtt acgcgcagcg tgaccgctac
     3121 acttgccagc gccctagcgc ccgctccttt cgctttcttc ccttcctttc tcgccacgtt
     3181 cgccggcttt ccccgtcaag ctctaaatcg ggggctccct ttagggttcc gatttagtgc
     3241 tttacggcac ctcgacccca aaaaacttga ttagggtgat ggttcacgta gtgggccatc
     3301 gccctgatag acggtttttc gccctttgac gttggagtcc acgttcttta atagtggact
     3361 cttgttccaa actggaacaa cactcaaccc tatctcggtc tattcttttg atttataagg
     3421 gattttgccg atttcggcct attggttaaa aaatgagctg atttaacaaa aatttaacgc
     3481 gaattaattc tgtggaatgt gtgtcagtta gggtgtggaa agtccccagg ctccccagca
     3541 ggcagaagta tgcaaagcat gcatctcaat tagtcagcaa ccaggtgtgg aaagtcccca
     3601 ggctccccag caggcagaag tatgcaaagc atgcatctca attagtcagc aaccatagtc
     3661 ccgcccctaa ctccgcccat cccgccccta actccgccca gttccgccca ttctccgccc
     3721 catggctgac taattttttt tatttatgca gaggccgagg ccgcctctgc ctctgagcta
     3781 ttccagaagt agtgaggagg cttttttgga ggcctaggct tttgcaaaaa gctcccggga
     3841 gcttgtatat ccattttcgg atctgatcaa gagacaggat gaggatcgtt tcgcatgatt
     3901 gaacaagatg gattgcacgc aggttctccg gccgcttggg tggagaggct attcggctat
     3961 gactgggcac aacagacaat cggctgctct gatgccgccg tgttccggct gtcagcgcag
     4021 gggcgcccgg ttctttttgt caagaccgac ctgtccggtg ccctgaatga actgcaggac
     4081 gaggcagcgc ggctatcgtg gctggccacg acgggcgttc cttgcgcagc tgtgctcgac
     4141 gttgtcactg aagcgggaag ggactggctg ctattgggcg aagtgccggg gcaggatctc
     4201 ctgtcatctc accttgctcc tgccgagaaa gtatccatca tggctgatgc aatgcggcgg
     4261 ctgcatacgc ttgatccggc tacctgccca ttcgaccacc aagcgaaaca tcgcatcgag
     4321 cgagcacgta ctcggatgga agccggtctt gtcgatcagg atgatctgga cgaagagcat
     4381 caggggctcg cgccagccga actgttcgcc aggctcaagg cgcgcatgcc cgacggcgag
     4441 gatctcgtcg tgacccatgg cgatgcctgc ttgccgaata tcatggtgga aaatggccgc
     4501 ttttctggat tcatcgactg tggccggctg ggtgtggcgg accgctatca ggacatagcg
     4561 ttggctaccc gtgatattgc tgaagagctt ggcggcgaat gggctgaccg cttcctcgtg
     4621 ctttacggta tcgccgctcc cgattcgcag cgcatcgcct tctatcgcct tcttgacgag
     4681 ttcttctgag cgggactctg gggttcgcga aatgaccgac caagcgacgc ccaacctgcc
     4741 atcacgagat ttcgattcca ccgccgcctt ctatgaaagg ttgggcttcg gaatcgtttt
     4801 ccgggacgcc ggctggatga tcctccagcg cggggatctc atgctggagt tcttcgccca
     4861 ccccaacttg tttattgcag cttataatgg ttacaaataa agcaatagca tcacaaattt
     4921 cacaaataaa gcattttttt cactgcattc tagttgtggt ttgtccaaac tcatcaatgt
     4981 atcttatcat gtctgtatac cgtcgacctc tagctagagc ttggcgtaat catggtcata
     5041 gctgtttcct gtgtgaaatt gttatccgct cacaattcca cacaacatac gagccggaag
     5101 cataaagtgt aaagcctggg gtgcctaatg agtgagctaa ctcacattaa ttgcgttgcg
     5161 ctcactgccc gctttccagt cgggaaacct gtcgtgccag ctgcattaat gaatcggcca
     5221 acgcgcgggg agaggcggtt tgcgtattgg gcgctcttcc gcttcctcgc tcactgactc
     5281 gctgcgctcg gtcgttcggc tgcggcgagc ggtatcagct cactcaaagg cggtaatacg
     5341 gttatccaca gaatcagggg ataacgcagg aaagaacatg tgagcaaaag gccagcaaaa
     5401 ggccaggaac cgtaaaaagg ccgcgttgct ggcgtttttc cataggctcc gcccccctga
     5461 cgagcatcac aaaaatcgac gctcaagtca gaggtggcga aacccgacag gactataaag
     5521 ataccaggcg tttccccctg gaagctccct cgtgcgctct cctgttccga ccctgccgct
     5581 taccggatac ctgtccgcct ttctcccttc gggaagcgtg gcgctttctc atagctcacg
     5641 ctgtaggtat ctcagttcgg tgtaggtcgt tcgctccaag ctgggctgtg tgcacgaacc
     5701 ccccgttcag cccgaccgct gcgccttatc cggtaactat cgtcttgagt ccaacccggt
     5761 aagacacgac ttatcgccac tggcagcagc cactggtaac aggattagca gagcgaggta
     5821 tgtaggcggt gctacagagt tcttgaagtg gtggcctaac tacggctaca ctagaagaac
     5881 agtatttggt atctgcgctc tgctgaagcc agttaccttc ggaaaaagag ttggtagctc
     5941 ttgatccggc aaacaaacca ccgctggtag cggtggtttt tttgtttgca agcagcagat
     6001 tacgcgcaga aaaaaaggat ctcaagaaga tcctttgatc ttttctacgg ggtctgacgc
     6061 tcagtggaac gaaaactcac gttaagggat tttggtcatg agattatcaa aaaggatctt
     6121 cacctagatc cttttaaatt aaaaatgaag ttttaaatca atctaaagta tatatgagta
     6181 aacttggtct gacagttacc aatgcttaat cagtgaggca cctatctcag cgatctgtct
     6241 atttcgttca tccatagttg cctgactccc cgtcgtgtag ataactacga tacgggaggg
     6301 cttaccatct ggccccagtg ctgcaatgat accgcgagac ccacgctcac cggctccaga
     6361 tttatcagca ataaaccagc cagccggaag ggccgagcgc agaagtggtc ctgcaacttt
     6421 atccgcctcc atccagtcta ttaattgttg ccgggaagct agagtaagta gttcgccagt
     6481 taatagtttg cgcaacgttg ttgccattgc tacaggcatc gtggtgtcac gctcgtcgtt
     6541 tggtatggct tcattcagct ccggttccca acgatcaagg cgagttacat gatcccccat
     6601 gttgtgcaaa aaagcggtta gctccttcgg tcctccgatc gttgtcagaa gtaagttggc
     6661 cgcagtgtta tcactcatgg ttatggcagc actgcataat tctcttactg tcatgccatc
     6721 cgtaagatgc ttttctgtga ctggtgagta ctcaaccaag tcattctgag aatagtgtat
     6781 gcggcgaccg agttgctctt gcccggcgtc aatacgggat aataccgcgc cacatagcag
     6841 aactttaaaa gtgctcatca ttggaaaacg ttcttcgggg cgaaaactct caaggatctt
     6901 accgctgttg agatccagtt cgatgtaacc cactcgtgca cccaactgat cttcagcatc
     6961 ttttactttc accagcgttt ctgggtgagc aaaaacagga aggcaaaatg ccgcaaaaaa
     7021 gggaataagg gcgacacgga aatgttgaat actcatactc ttcctttttc aatattattg
     7081 aagcatttat cagggttatt gtctcatgag cggatacata tttgaatgta tttagaaaaa
     7141 taaacaaata ggggttccgc gcacatttcc ccgaaaagtg ccacctgacg
//
