LOCUS       sequence-199327-        7083 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Anion channelrhodopsin from Rhodomonas sp. (CCMP768) for expression 
            in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7083)
  AUTHORS   Govorunova EG, Sineshchekov OA, Rodarte EM, Janz R, Morelle O, 
            Melkonian M, Wong GK, Spudich JL
  TITLE     The Expanding Family of Natural Anion Channelrhodopsins Reveals 
            Large Variations in Kinetics, Conductance, and Spectral Sensitivity.
  JOURNAL   Sci Rep. 2017 Mar 3;7:43358. doi: 10.1038/srep43358.
  PUBMED    28256618
REFERENCE   2  (bases 1 to 7083)
  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: "2017-03-3"; volume: "7:43358. doi"; pages: " 
            10.1038/srep43358"
COMMENT     Sequence Label: pcDNA3.1-R1ACR_447-EYFP
FEATURES             Location/Qualifiers
     source          1..7083
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     complement(75..95)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        80..409
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      260..395
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     322..341
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             476..1270
                     /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(530..549)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     1140..1159
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    1446..1567
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(1483..1502)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     1537..1556
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(1616..1632)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(1616..1632)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(1629..1651)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    1640..1656
                     /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(1664..1670,1671..1688,1689..1694))
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    1709..1730
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(1847..1864)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(2018..2606)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(2098..2117)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(join(2777..3568,3569..3637))
                     /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     3400..3419
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(3638..3742)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(3817..3836)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        4008..4387
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        4388..4591
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     4542..4562
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     4636..4655
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        4636..4654
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             join(5608..5610,5611..6324)
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(5650..5671)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(5911..5930)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     6258..6279
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             6354..6395
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             6405..6422
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(6445..6462)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    6451..6675
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      join(6721..7083,1..66)
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6808..6827)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     7018..7039
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
ORIGIN
        1 ggattttgcc gatttcggcc tattggttaa aaaatgagct gatttaacaa aaatttaacg
       61 cgaattaatt ctgtggaatg tgtgtcagtt agggtgtgga aagtccccag gctccccagc
      121 aggcagaagt atgcaaagca tgcatctcaa ttagtcagca accaggtgtg gaaagtcccc
      181 aggctcccca gcaggcagaa gtatgcaaag catgcatctc aattagtcag caaccatagt
      241 cccgccccta actccgccca tcccgcccct aactccgccc agttccgccc attctccgcc
      301 ccatggctga ctaatttttt ttatttatgc agaggccgag gccgcctctg cctctgagct
      361 attccagaag tagtgaggag gcttttttgg aggcctaggc ttttgcaaaa agctcccggg
      421 agcttgtata tccattttcg gatctgatca agagacagga tgaggatcgt ttcgcatgat
      481 tgaacaagat ggattgcacg caggttctcc ggccgcttgg gtggagaggc tattcggcta
      541 tgactgggca caacagacaa tcggctgctc tgatgccgcc gtgttccggc tgtcagcgca
      601 ggggcgcccg gttctttttg tcaagaccga cctgtccggt gccctgaatg aactgcagga
      661 cgaggcagcg cggctatcgt ggctggccac gacgggcgtt ccttgcgcag ctgtgctcga
      721 cgttgtcact gaagcgggaa gggactggct gctattgggc gaagtgccgg ggcaggatct
      781 cctgtcatct caccttgctc ctgccgagaa agtatccatc atggctgatg caatgcggcg
      841 gctgcatacg cttgatccgg ctacctgccc attcgaccac caagcgaaac atcgcatcga
      901 gcgagcacgt actcggatgg aagccggtct tgtcgatcag gatgatctgg acgaagagca
      961 tcaggggctc gcgccagccg aactgttcgc caggctcaag gcgcgcatgc ccgacggcga
     1021 ggatctcgtc gtgacccatg gcgatgcctg cttgccgaat atcatggtgg aaaatggccg
     1081 cttttctgga ttcatcgact gtggccggct gggtgtggcg gaccgctatc aggacatagc
     1141 gttggctacc cgtgatattg ctgaagagct tggcggcgaa tgggctgacc gcttcctcgt
     1201 gctttacggt atcgccgctc ccgattcgca gcgcatcgcc ttctatcgcc ttcttgacga
     1261 gttcttctga gcgggactct ggggttcgcg aaatgaccga ccaagcgacg cccaacctgc
     1321 catcacgaga tttcgattcc accgccgcct tctatgaaag gttgggcttc ggaatcgttt
     1381 tccgggacgc cggctggatg atcctccagc gcggggatct catgctggag ttcttcgccc
     1441 accccaactt gtttattgca gcttataatg gttacaaata aagcaatagc atcacaaatt
     1501 tcacaaataa agcatttttt tcactgcatt ctagttgtgg tttgtccaaa ctcatcaatg
     1561 tatcttatca tgtctgtata ccgtcgacct ctagctagag cttggcgtaa tcatggtcat
     1621 agctgtttcc tgtgtgaaat tgttatccgc tcacaattcc acacaacata cgagccggaa
     1681 gcataaagtg taaagcctgg ggtgcctaat gagtgagcta actcacatta attgcgttgc
     1741 gctcactgcc cgctttccag tcgggaaacc tgtcgtgcca gctgcattaa tgaatcggcc
     1801 aacgcgcggg gagaggcggt ttgcgtattg ggcgctcttc cgcttcctcg ctcactgact
     1861 cgctgcgctc ggtcgttcgg ctgcggcgag cggtatcagc tcactcaaag gcggtaatac
     1921 ggttatccac agaatcaggg gataacgcag gaaagaacat gtgagcaaaa ggccagcaaa
     1981 aggccaggaa ccgtaaaaag gccgcgttgc tggcgttttt ccataggctc cgcccccctg
     2041 acgagcatca caaaaatcga cgctcaagtc agaggtggcg aaacccgaca ggactataaa
     2101 gataccaggc gtttccccct ggaagctccc tcgtgcgctc tcctgttccg accctgccgc
     2161 ttaccggata cctgtccgcc tttctccctt cgggaagcgt ggcgctttct catagctcac
     2221 gctgtaggta tctcagttcg gtgtaggtcg ttcgctccaa gctgggctgt gtgcacgaac
     2281 cccccgttca gcccgaccgc tgcgccttat ccggtaacta tcgtcttgag tccaacccgg
     2341 taagacacga cttatcgcca ctggcagcag ccactggtaa caggattagc agagcgaggt
     2401 atgtaggcgg tgctacagag ttcttgaagt ggtggcctaa ctacggctac actagaagaa
     2461 cagtatttgg tatctgcgct ctgctgaagc cagttacctt cggaaaaaga gttggtagct
     2521 cttgatccgg caaacaaacc accgctggta gcggtggttt ttttgtttgc aagcagcaga
     2581 ttacgcgcag aaaaaaagga tctcaagaag atcctttgat cttttctacg gggtctgacg
     2641 ctcagtggaa cgaaaactca cgttaaggga ttttggtcat gagattatca aaaaggatct
     2701 tcacctagat ccttttaaat taaaaatgaa gttttaaatc aatctaaagt atatatgagt
     2761 aaacttggtc tgacagttac caatgcttaa tcagtgaggc acctatctca gcgatctgtc
     2821 tatttcgttc atccatagtt gcctgactcc ccgtcgtgta gataactacg atacgggagg
     2881 gcttaccatc tggccccagt gctgcaatga taccgcgaga cccacgctca ccggctccag
     2941 atttatcagc aataaaccag ccagccggaa gggccgagcg cagaagtggt cctgcaactt
     3001 tatccgcctc catccagtct attaattgtt gccgggaagc tagagtaagt agttcgccag
     3061 ttaatagttt gcgcaacgtt gttgccattg ctacaggcat cgtggtgtca cgctcgtcgt
     3121 ttggtatggc ttcattcagc tccggttccc aacgatcaag gcgagttaca tgatccccca
     3181 tgttgtgcaa aaaagcggtt agctccttcg gtcctccgat cgttgtcaga agtaagttgg
     3241 ccgcagtgtt atcactcatg gttatggcag cactgcataa ttctcttact gtcatgccat
     3301 ccgtaagatg cttttctgtg actggtgagt actcaaccaa gtcattctga gaatagtgta
     3361 tgcggcgacc gagttgctct tgcccggcgt caatacggga taataccgcg ccacatagca
     3421 gaactttaaa agtgctcatc attggaaaac gttcttcggg gcgaaaactc tcaaggatct
     3481 taccgctgtt gagatccagt tcgatgtaac ccactcgtgc acccaactga tcttcagcat
     3541 cttttacttt caccagcgtt tctgggtgag caaaaacagg aaggcaaaat gccgcaaaaa
     3601 agggaataag ggcgacacgg aaatgttgaa tactcatact cttccttttt caatattatt
     3661 gaagcattta tcagggttat tgtctcatga gcggatacat atttgaatgt atttagaaaa
     3721 ataaacaaat aggggttccg cgcacatttc cccgaaaagt gccacctgac gtcgacggat
     3781 cgggagatct cccgatcccc tatggtgcac tctcagtaca atctgctctg atgccgcata
     3841 gttaagccag tatctgctcc ctgcttgtgt gttggaggtc gctgagtagt gcgcgagcaa
     3901 aatttaagct acaacaaggc aaggcttgac cgacaattgc atgaagaatc tgcttagggt
     3961 taggcgtttt gcgctgcttc gcgatgtacg ggccagatat acgcgttgac attgattatt
     4021 gactagttat taatagtaat caattacggg gtcattagtt catagcccat atatggagtt
     4081 ccgcgttaca taacttacgg taaatggccc gcctggctga ccgcccaacg acccccgccc
     4141 attgacgtca ataatgacgt atgttcccat agtaacgcca atagggactt tccattgacg
     4201 tcaatgggtg gagtatttac ggtaaactgc ccacttggca gtacatcaag tgtatcatat
     4261 gccaagtacg ccccctattg acgtcaatga cggtaaatgg cccgcctggc attatgccca
     4321 gtacatgacc ttatgggact ttcctacttg gcagtacatc tacgtattag tcatcgctat
     4381 taccatggtg atgcggtttt ggcagtacat caatgggcgt ggatagcggt ttgactcacg
     4441 gggatttcca agtctccacc ccattgacgt caatgggagt ttgttttggc accaaaatca
     4501 acgggacttt ccaaaatgtc gtaacaactc cgccccattg acgcaaatgg gcggtaggcg
     4561 tgtacggtgg gaggtctata taagcagagc tctctggcta actagagaac ccactgctta
     4621 ctggcttatc gaaattaata cgactcacta tagggagacc caagctggct agttaagctt
     4681 ggtaccgagc tcggatccat ggtgtggtcc catgtggacg agcctgccct gggaccctat
     4741 ggcagctggg aagccgccaa ccccgattgt gtggccctgg gcctgaccac ctccaagaac
     4801 accaagatga tcctcctgct gttcgccagc atttgcgcct gtcaggtgct gttcatggcc
     4861 agcagggccc ctaggattac atgggaggcc gtctacctgc ctctgatcga gaccatcctg
     4921 tacgctatgg ccagcaccgg caatggccag ctgaggctgg gagacggcag ggtgctgcct
     4981 atcgccaggt actgctcctg gctgatcacc tgccccatca tgctgttcca ggtggtggga
     5041 ctctacgatc tgaagctctg gggaatctcc tgcaagcaca tggtgatggc cgccgccctg
     5101 atcagaaccg tctttggaat cggcgcttcc gtgacactgg atgaccagat gaggtgggtg
     5161 cagttctttc tgtcctacgc cttcttcttc tttgagctgt actgcgccta catgatcttc
     5221 ggcgctgccc tgaagcagtt cggcgaagtg agaaccaccc tgaatagcat cgtgtactcc
     5281 aggatcatgc tgctgaggtt cattttcttc ctgagctgga acagcttcgg catcatctgg
     5341 ctgctgagca gcaccggcct gtgcatgatt tccgaggacg tgtccgtgct ggcctacctg
     5401 gccgctgaca tgctgtgcaa gaacacctac ggcatcatca acgccaacac cctgttcaga
     5461 gtcctggacg gcaagtggac ccccgagaca ctggaggaag ccgcctccgg cgtgaagaag
     5521 gccaacccca accacgtggg cgagacatac ctggagagga tgatggccgg cattaggggc
     5581 agcgctaagc ccgaggaggc ggccgccgtg agcaagggcg aggagctgtt caccggggtg
     5641 gtgcccatcc tggtcgagct ggacggcgac gtaaacggcc acaagttcag cgtgtccggc
     5701 gagggcgagg gcgatgccac ctacggcaag ctgaccctga agttcatctg caccaccggc
     5761 aagctgcccg tgccctggcc caccctcgtg accaccttcg gctacggcct gcagtgcttc
     5821 gcccgctacc ccgaccacat gaagcagcac gacttcttca agtccgccat gcccgaaggc
     5881 tacgtccagg agcgcaccat cttcttcaag gacgacggca actacaagac ccgcgccgag
     5941 gtgaagttcg agggcgacac cctggtgaac cgcatcgagc tgaagggcat cgacttcaag
     6001 gaggacggca acatcctggg gcacaagctg gagtacaact acaacagcca caacgtctat
     6061 atcatggccg acaagcagaa gaacggcatc aaggtgaact tcaagatccg ccacaacatc
     6121 gaggacggca gcgtgcagct cgccgaccac taccagcaga acacccccat cggcgacggc
     6181 cccgtgctgc tgcccgacaa ccactacctg agctaccagt ccgccctgag caaagacccc
     6241 aacgagaagc gcgatcacat ggtcctgctg gagttcgtga ccgccgccgg gatcactctc
     6301 ggcatggacg agctgtacaa gtaacatatg ctctagaggg cccgcggttc gaaggtaagc
     6361 ctatccctaa ccctctcctc ggtctcgatt ctacgcgtac cggtcatcat caccatcacc
     6421 attgagttta aacccgctga tcagcctcga ctgtgccttc tagttgccag ccatctgttg
     6481 tttgcccctc ccccgtgcct tccttgaccc tggaaggtgc cactcccact gtcctttcct
     6541 aataaaatga ggaaattgca tcgcattgtc tgagtaggtg tcattctatt ctggggggtg
     6601 gggtggggca ggacagcaag ggggaggatt gggaagacaa tagcaggcat gctggggatg
     6661 cggtgggctc tatggcttct gaggcggaaa gaaccagctg gggctctagg gggtatcccc
     6721 acgcgccctg tagcggcgca ttaagcgcgg cgggtgtggt ggttacgcgc agcgtgaccg
     6781 ctacacttgc cagcgcccta gcgcccgctc ctttcgcttt cttcccttcc tttctcgcca
     6841 cgttcgccgg ctttccccgt caagctctaa atcgggggct ccctttaggg ttccgattta
     6901 gtgctttacg gcacctcgac cccaaaaaac ttgattaggg tgatggttca cgtagtgggc
     6961 catcgccctg atagacggtt tttcgccctt tgacgttgga gtccacgttc tttaatagtg
     7021 gactcttgtt ccaaactgga acaacactca accctatctc ggtctattct tttgatttat
     7081 aag
//
