LOCUS       sequence-182725-        5659 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  slow cycling (open for minutes) step-function artificial anion 
            conducting channelrhodopsin (aACR). High light sensitivity. 
            Activation with blue to green light, inactivation max with 605 nm 
            (accelarates closure to ms). Codon optimized for mammalian 
            expression..
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 5659)
  AUTHORS   Wietek J, Rodriguez-Rozada S, Tutas J, Tenedini F, Grimm C, Oertner 
            TG, Soba P, Hegemann P, Wiegert JS
  TITLE     Anion-conducting channelrhodopsins with tuned spectra and modified 
            kinetics engineered for optogenetic manipulation of behavior.
  JOURNAL   Sci Rep. 2017 Nov 2;7(1):14957. doi: 10.1038/s41598-017-14330-y.
  PUBMED    29097684
REFERENCE   2  (bases 1 to 5659)
  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: "Sci Rep."; 
            date: "2017-11-2"; volume: "7(1):14957. doi"; pages: " 
            10.1038/s41598-017-14330-y"
COMMENT     Sequence Label: p-mCherry-N1-iChloC_CA
FEATURES             Location/Qualifiers
     source          1..5659
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     366..385
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     enhancer        640..943
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        944..1147
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     1098..1118
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     misc_feature    1170..1250
                     /label=MCS
                     /note="multiple cloning site"
     regulatory      1251..1260
                     /label=Kozak sequence
                     /note="vertebrate consensus sequence for strong initiation 
                     of translation (Kozak, 1987)"
                     /regulatory_class="other"
     CDS             1257..2183
                     /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
                     NGAQTASNVLQWLAAGFSILLLMFYAYQTWKSTCGWEQIYVCAIRMVKVILEFFFSFKN
                     PSMLYLATGHRVQWLRYAEWLLTAPVILIHLSNLTGLSNDYSRRTMGLLVSNIGCIVWG
                     ATSAMATGYVKVIFFCLGLCYGANTFFHAAKAYIEGYHTVPKGRCRQVVTGMAWLFFVS
                     WGMFPILFILGPEGFGVLSVYGSTVGHTIIDLMSKNCWGLLGHYLRVLIHEHILIHGDI
                     RKTTKLNIGGTEIEVETLVEDEAEAGAV"
     CDS             2196..2903
                     /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(2338..2356)
                     /label=mCherry-R
                     /note="mCherry, reverse primer"
     primer_bind     2606..2625
                     /label=mCherry-F
                     /note="mCherry, forward primer"
     polyA_signal    3026..3147
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(3063..3082)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     3117..3136
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     rep_origin      complement(3154..3609)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(3291..3312)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     3503..3522
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     promoter        3636..3740
                     /gene="bla"
                     /label=AmpR promoter
     promoter        3742..4099
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     enhancer        3742..3933
                     /label=SV40 enhancer
                     /note="enhancer for the SV40 early promoter (Herr, 1993)"
     primer_bind     complement(3765..3785)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      3950..4085
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     CDS             4134..4928
                     /codon_start=1
                     /product="aminoglycoside phosphotransferase"
                     /label=NeoR/KanR
                     /note="confers resistance to neomycin, kanamycin, and G418 
                     (Geneticin(R))"
                     /translation="MIEQDGLHAGSPAAWVERLFGYDWAQQTIGCSDAAVFRLSAQGRP
                     VLFVKTDLSGALNELQDEAARLSWLATTGVPCAAVLDVVTEAGRDWLLLGEVPGQDLLS
                     SHLAPAEKVSIMADAMRRLHTLDPATCPFDHQAKHRIERARTRMEAGLVDQDDLDEEHQ
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     CDS             4134..4928
                     /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
                     GLAPAELFARLKASMPDGEDLVVTHGDACLPNIMVENGRFSGFIDCGRLGVADRYQDIA
                     LATRDIAEELGGEWADRFLVLYGIAAPDSQRIAFYRLLDEFF"
     primer_bind     complement(4188..4207)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     4798..4817
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     complement(5116..5135)
                     /label=TK-pA-R
                     /note="Thymidine kinase polyA, reverse primer"
     polyA_signal    5160..5207
                     /label=HSV TK poly(A) signal
                     /note="herpes simplex virus thymidine kinase 
                     polyadenylation signal (Cole and Stacy, 1985)"
     rep_origin      join(5536..5659,1..465)
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
ORIGIN
        1 agagcgcaga taccaaatac tgttcttcta gtgtagccgt agttaggcca ccacttcaag
       61 aactctgtag caccgcctac atacctcgct ctgctaatcc tgttaccagt ggctgctgcc
      121 agtggcgata agtcgtgtct taccgggttg gactcaagac gatagttacc ggataaggcg
      181 cagcggtcgg gctgaacggg gggttcgtgc acacagccca gcttggagcg aacgacctac
      241 accgaactga gatacctaca gcgtgagcta tgagaaagcg ccacgcttcc cgaagggaga
      301 aaggcggaca ggtatccggt aagcggcagg gtcggaacag gagagcgcac gagggagctt
      361 ccagggggaa acgcctggta tctttatagt cctgtcgggt ttcgccacct ctgacttgag
      421 cgtcgatttt tgtgatgctc gtcagggggg cggagcctat ggaaaaacgc cagcaacgcg
      481 gcctttttac ggttcctggc cttttgctgg ccttttgctc acatgttctt tcctgcgtta
      541 tcccctgatt ctgtggataa ccgtattacc gccatgcatt agttattaat agtaatcaat
      601 tacggggtca ttagttcata gcccatatat ggagttccgc gttacataac ttacggtaaa
      661 tggcccgcct ggctgaccgc ccaacgaccc ccgcccattg acgtcaataa tgacgtatgt
      721 tcccatagta acgccaatag ggactttcca ttgacgtcaa tgggtggagt atttacggta
      781 aactgcccac ttggcagtac atcaagtgta tcatatgcca agtacgcccc ctattgacgt
      841 caatgacggt aaatggcccg cctggcatta tgcccagtac atgaccttat gggactttcc
      901 tacttggcag tacatctacg tattagtcat cgctattacc atggtgatgc ggttttggca
      961 gtacatcaat gggcgtggat agcggtttga ctcacgggga tttccaagtc tccaccccat
     1021 tgacgtcaat gggagtttgt tttggcacca aaatcaacgg gactttccaa aatgtcgtaa
     1081 caactccgcc ccattgacgc aaatgggcgg taggcgtgta cggtgggagg tctatataag
     1141 cagagctggt ttagtgaacc gtcagatccg ctagcgctac cggactcaga tctcgagctc
     1201 aagcttcgaa ttctgcagtc gacggtaccg cgggcccggg atccaccggt gccaccatgg
     1261 actatggcgg cgctttgtct gccgtcggac gcgaactttt gttcgttact aatcctgtgg
     1321 tggtgaacgg gtccgtcctg gtccctgagg atcaatgtta ctgtgccgga tggattgaat
     1381 ctcgcggcac gaacggcgct cagaccgcgt caaatgtcct gcagtggctt gcagcaggat
     1441 tcagcatttt gctgctgatg ttctatgcct accaaacctg gaaatctaca tgcggctggg
     1501 agcagatcta tgtgtgcgcc attagaatgg ttaaggtgat tctcgagttc ttttttagct
     1561 ttaagaatcc ctctatgctc taccttgcca caggacaccg ggtgcagtgg ctgcgctatg
     1621 cagagtggct gctcactgct cctgtcatcc ttatccacct gagcaacctc accggcctga
     1681 gcaacgacta cagcaggaga accatgggac tccttgtctc aaacatcggg tgtatcgtgt
     1741 ggggggctac cagcgccatg gcaaccggct atgttaaagt catcttcttt tgtcttggat
     1801 tgtgctatgg cgcgaacaca ttttttcacg ccgccaaagc atatatcgag ggttatcata
     1861 ctgtgccaaa gggtcggtgc cgccaggtcg tgaccggcat ggcatggctg tttttcgtga
     1921 gctggggtat gttcccaatt ctcttcattt tggggcccga aggttttggc gtcctgagcg
     1981 tctatggctc caccgtaggt cacacgatta ttgatctgat gagtaaaaat tgttgggggt
     2041 tgttgggaca ctacctgcgc gtcctgatcc acgagcacat attgattcac ggagatatcc
     2101 gcaaaaccac caaactgaac atcggcggaa cggagatcga ggtcgagact ctcgtcgaag
     2161 acgaagccga ggccggagcc gtgccagcgg ccgccgtgag caagggcgag gaggataaca
     2221 tggccatcat caaggagttc atgcgcttca aggtgcacat ggagggctcc gtgaacggcc
     2281 acgagttcga gatcgagggc gagggcgagg gccgccccta cgagggcacc cagaccgcca
     2341 agctgaaggt gaccaagggt ggccccctgc ccttcgcctg ggacatcctg tcccctcagt
     2401 tcatgtacgg ctccaaggcc tacgtgaagc accccgccga catccccgac tacttgaagc
     2461 tgtccttccc cgagggcttc aagtgggagc gcgtgatgaa cttcgaggac ggcggcgtgg
     2521 tgaccgtgac ccaggactcc tccctgcagg acggcgagtt catctacaag gtgaagctgc
     2581 gcggcaccaa cttcccctcc gacggccccg taatgcagaa gaagaccatg ggctgggagg
     2641 cctcctccga gcggatgtac cccgaggacg gcgccctgaa gggcgagatc aagcagaggc
     2701 tgaagctgaa ggacggcggc cactacgacg ctgaggtcaa gaccacctac aaggccaaga
     2761 agcccgtgca gctgcccggc gcctacaacg tcaacatcaa gttggacatc acctcccaca
     2821 acgaggacta caccatcgtg gaacagtacg aacgcgccga gggccgccac tccaccggcg
     2881 gcatggacga gctgtacaag taaagcggcc gcgactctag atcataatca gccataccac
     2941 atttgtagag gttttacttg ctttaaaaaa cctcccacac ctccccctga acctgaaaca
     3001 taaaatgaat gcaattgttg ttgttaactt gtttattgca gcttataatg gttacaaata
     3061 aagcaatagc atcacaaatt tcacaaataa agcatttttt tcactgcatt ctagttgtgg
     3121 tttgtccaaa ctcatcaatg tatcttaagg cgtaaattgt aagcgttaat attttgttaa
     3181 aattcgcgtt aaatttttgt taaatcagct cattttttaa ccaataggcc gaaatcggca
     3241 aaatccctta taaatcaaaa gaatagaccg agatagggtt gagtgttgtt ccagtttgga
     3301 acaagagtcc actattaaag aacgtggact ccaacgtcaa agggcgaaaa accgtctatc
     3361 agggcgatgg cccactacgt gaaccatcac cctaatcaag ttttttgggg tcgaggtgcc
     3421 gtaaagcact aaatcggaac cctaaaggga gcccccgatt tagagcttga cggggaaagc
     3481 cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg agcgggcgct agggcgctgg
     3541 caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc cgcgcttaat gcgccgctac
     3601 agggcgcgtc aggtggcact tttcggggaa atgtgcgcgg aacccctatt tgtttatttt
     3661 tctaaataca ttcaaatatg tatccgctca tgagacaata accctgataa atgcttcaat
     3721 aatattgaaa aaggaagagt cctgaggcgg aaagaaccag ctgtggaatg tgtgtcagtt
     3781 agggtgtgga aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa
     3841 ttagtcagca accaggtgtg gaaagtcccc aggctcccca gcaggcagaa gtatgcaaag
     3901 catgcatctc aattagtcag caaccatagt cccgccccta actccgccca tcccgcccct
     3961 aactccgccc agttccgccc attctccgcc ccatggctga ctaatttttt ttatttatgc
     4021 agaggccgag gccgcctcgg cctctgagct attccagaag tagtgaggag gcttttttgg
     4081 aggcctaggc ttttgcaaag atcgatcaag agacaggatg aggatcgttt cgcatgattg
     4141 aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta ttcggctatg
     4201 actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg tcagcgcagg
     4261 ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa ctgcaagacg
     4321 aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct gtgctcgacg
     4381 ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg caggatctcc
     4441 tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca atgcggcggc
     4501 tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat cgcatcgagc
     4561 gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac gaagagcatc
     4621 aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gagcatgccc gacggcgagg
     4681 atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa aatggccgct
     4741 tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag gacatagcgt
     4801 tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc ttcctcgtgc
     4861 tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt cttgacgagt
     4921 tcttctgagc gggactctgg ggttcgaaat gaccgaccaa gcgacgccca acctgccatc
     4981 acgagatttc gattccaccg ccgccttcta tgaaaggttg ggcttcggaa tcgttttccg
     5041 ggacgccggc tggatgatcc tccagcgcgg ggatctcatg ctggagttct tcgcccaccc
     5101 tagggggagg ctaactgaaa cacggaagga gacaataccg gaaggaaccc gcgctatgac
     5161 ggcaataaaa agacagaata aaacgcacgg tgttgggtcg tttgttcata aacgcggggt
     5221 tcggtcccag ggctggcact ctgtcgatac cccaccgaga ccccattggg gccaatacgc
     5281 ccgcgtttct tccttttccc caccccaccc cccaagttcg ggtgaaggcc cagggctcgc
     5341 agccaacgtc ggggcggcag gccctgccat agcctcaggt tactcatata tactttagat
     5401 tgatttaaaa cttcattttt aatttaaaag gatctaggtg aagatccttt ttgataatct
     5461 catgaccaaa atcccttaac gtgagttttc gttccactga gcgtcagacc ccgtagaaaa
     5521 gatcaaagga tcttcttgag atcctttttt tctgcgcgta atctgctgct tgcaaacaaa
     5581 aaaaccaccg ctaccagcgg tggtttgttt gccggatcaa gagctaccaa ctctttttcc
     5641 gaaggtaact ggcttcagc
//
