LOCUS       sequence-199236-        7086 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Anion channelrhodopsin from Proteomonas sulcata (CCMP704) for 
            expression in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7086)
  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 7086)
  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-PsuACR_433-EYFP
FEATURES             Location/Qualifiers
     source          1..7086
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     162..179
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(202..219)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(229..270)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             complement(join(300..1013,1014..1016))
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(345..366)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     694..713
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     953..974
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(1972..1991)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(1973..1991)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(2036..2239)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2065..2085)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        2240..2619
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     2791..2810
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        2885..2989
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(2990..3058,3059..3850)
                     /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(3208..3227)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4021..4609
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     4510..4529
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     4763..4780
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    4897..4918
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(4933..4938,4939..4956,4957..4963)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    4971..4987
                     /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)."
     primer_bind     4976..4998
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     4995..5011
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     4995..5011
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    5060..5181
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5071..5090)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5125..5144
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(5357..6151)
                     /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(5468..5487)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     6078..6097
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(6218..6547)
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      6232..6367
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(6286..6305)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     primer_bind     6532..6552
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(6561..6989)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(6671..6692)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     6883..6902
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    join(7035..7086,1..173)
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
ORIGIN
        1 ccccttgctg tcctgcccca ccccaccccc cagaatagaa tgacacctac tcagacaatg
       61 cgatgcaatt tcctcatttt attaggaaag gacagtggga gtggcacctt ccagggtcaa
      121 ggaaggcacg ggggaggggc aaacaacaga tggctggcaa ctagaaggca cagtcgaggc
      181 tgatcagcgg gtttaaactc aatggtgatg gtgatgatga ccggtacgcg tagaatcgag
      241 accgaggaga gggttaggga taggcttacc ttcgaaccgc gggccctcta gagcatatgt
      301 tacttgtaca gctcgtccat gccgagagtg atcccggcgg cggtcacgaa ctccagcagg
      361 accatgtgat cgcgcttctc gttggggtct ttgctcaggg cggactggta gctcaggtag
      421 tggttgtcgg gcagcagcac ggggccgtcg ccgatggggg tgttctgctg gtagtggtcg
      481 gcgagctgca cgctgccgtc ctcgatgttg tggcggatct tgaagttcac cttgatgccg
      541 ttcttctgct tgtcggccat gatatagacg ttgtggctgt tgtagttgta ctccagcttg
      601 tgccccagga tgttgccgtc ctccttgaag tcgatgccct tcagctcgat gcggttcacc
      661 agggtgtcgc cctcgaactt cacctcggcg cgggtcttgt agttgccgtc gtccttgaag
      721 aagatggtgc gctcctggac gtagccttcg ggcatggcgg acttgaagaa gtcgtgctgc
      781 ttcatgtggt cggggtagcg ggcgaagcac tgcaggccgt agccgaaggt ggtcacgagg
      841 gtgggccagg gcacgggcag cttgccggtg gtgcagatga acttcagggt cagcttgccg
      901 taggtggcat cgccctcgcc ctcgccggac acgctgaact tgtggccgtt tacgtcgccg
      961 tccagctcga ccaggatggg caccaccccg gtgaacagct cctcgccctt gctcacggcg
     1021 gccgcgccat cgtgatcgtg gcagtcctgg gtgtccttca cgaagttgat gctctcggcc
     1081 aggccaatgc tcttgccggg gagggagtgt gtggcgctag cggagctcct agcctgctcc
     1141 tgatcggggg ggtagtggtg ctcgaagctc cagtcgccgt ccaggtctct ccagagggtc
     1201 caccacatca ccaggccaaa gatgttcttg gcgaagaagt cggacacgaa gtgcaggcag
     1261 gagatcacgt cttctctcag gctacaggtg gcctgggggg ataaaatcca cagcagaggg
     1321 tagatgctcc agctgaggct gaacaggtac ttcaggaagg tgaccctcca ggcgaccttc
     1381 tgggccaggg gtgtgtccac ggacttccac ttcctagcgg tctgggagaa gatatcgtgg
     1441 gtgatgccca tgatgcatcc gaagagcacc atggcaatgc cgaagtagat ccacttcagg
     1501 taggtgtggt tggtgagtgt ggcgttcagt cccatcacgt tcatgatcag atcggccacc
     1561 acctggaagg tgttcaggtt gaagtggccc agcttgcact tggccaggcc gttgatctgc
     1621 atcagcagag cgggacaggt acaaatccag gcggccatcc tggcataagg gatgatcctg
     1681 ccgtcggcca gctggatgaa gccattgccg gaagcggcca gggaatacag tgtcatctcg
     1741 caagcgggca ggtagatgct ttcccatccc actttgggcc actgtcccat gatccagatg
     1801 atctggcaca ggcacaccat gcagaagcac agctgggcgg ttttggtgtt ggggctggtg
     1861 taggacatct ccatgcactc ggggttctcc agggcccaca ggccgtactc gtccaccagc
     1921 tcagccatgg atccgagctc ggtaccaagc ttaactagcc agcttgggtc tccctatagt
     1981 gagtcgtatt aatttcgata agccagtaag cagtgggttc tctagttagc cagagagctc
     2041 tgcttatata gacctcccac cgtacacgcc taccgcccat ttgcgtcaat ggggcggagt
     2101 tgttacgaca ttttggaaag tcccgttgat tttggtgcca aaacaaactc ccattgacgt
     2161 caatggggtg gagacttgga aatccccgtg agtcaaaccg ctatccacgc ccattgatgt
     2221 actgccaaaa ccgcatcacc atggtaatag cgatgactaa tacgtagatg tactgccaag
     2281 taggaaagtc ccataaggtc atgtactggg cataatgcca ggcgggccat ttaccgtcat
     2341 tgacgtcaat agggggcgta cttggcatat gatacacttg atgtactgcc aagtgggcag
     2401 tttaccgtaa atactccacc cattgacgtc aatggaaagt ccctattggc gttactatgg
     2461 gaacatacgt cattattgac gtcaatgggc gggggtcgtt gggcggtcag ccaggcgggc
     2521 catttaccgt aagttatgta acgcggaact ccatatatgg gctatgaact aatgaccccg
     2581 taattgatta ctattaataa ctagtcaata atcaatgtca acgcgtatat ctggcccgta
     2641 catcgcgaag cagcgcaaaa cgcctaaccc taagcagatt cttcatgcaa ttgtcggtca
     2701 agccttgcct tgttgtagct taaattttgc tcgcgcacta ctcagcgacc tccaacacac
     2761 aagcagggag cagatactgg cttaactatg cggcatcaga gcagattgta ctgagagtgc
     2821 accatagggg atcgggagat ctcccgatcc gtcgacgtca ggtggcactt ttcggggaaa
     2881 tgtgcgcgga acccctattt gtttattttt ctaaatacat tcaaatatgt atccgctcat
     2941 gagacaataa ccctgataaa tgcttcaata atattgaaaa aggaagagta tgagtattca
     3001 acatttccgt gtcgccctta ttcccttttt tgcggcattt tgccttcctg tttttgctca
     3061 cccagaaacg ctggtgaaag taaaagatgc tgaagatcag ttgggtgcac gagtgggtta
     3121 catcgaactg gatctcaaca gcggtaagat ccttgagagt tttcgccccg aagaacgttt
     3181 tccaatgatg agcactttta aagttctgct atgtggcgcg gtattatccc gtattgacgc
     3241 cgggcaagag caactcggtc gccgcataca ctattctcag aatgacttgg ttgagtactc
     3301 accagtcaca gaaaagcatc ttacggatgg catgacagta agagaattat gcagtgctgc
     3361 cataaccatg agtgataaca ctgcggccaa cttacttctg acaacgatcg gaggaccgaa
     3421 ggagctaacc gcttttttgc acaacatggg ggatcatgta actcgccttg atcgttggga
     3481 accggagctg aatgaagcca taccaaacga cgagcgtgac accacgatgc ctgtagcaat
     3541 ggcaacaacg ttgcgcaaac tattaactgg cgaactactt actctagctt cccggcaaca
     3601 attaatagac tggatggagg cggataaagt tgcaggacca cttctgcgct cggcccttcc
     3661 ggctggctgg tttattgctg ataaatctgg agccggtgag cgtgggtctc gcggtatcat
     3721 tgcagcactg gggccagatg gtaagccctc ccgtatcgta gttatctaca cgacggggag
     3781 tcaggcaact atggatgaac gaaatagaca gatcgctgag ataggtgcct cactgattaa
     3841 gcattggtaa ctgtcagacc aagtttactc atatatactt tagattgatt taaaacttca
     3901 tttttaattt aaaaggatct aggtgaagat cctttttgat aatctcatga ccaaaatccc
     3961 ttaacgtgag ttttcgttcc actgagcgtc agaccccgta gaaaagatca aaggatcttc
     4021 ttgagatcct ttttttctgc gcgtaatctg ctgcttgcaa acaaaaaaac caccgctacc
     4081 agcggtggtt tgtttgccgg atcaagagct accaactctt tttccgaagg taactggctt
     4141 cagcagagcg cagataccaa atactgttct tctagtgtag ccgtagttag gccaccactt
     4201 caagaactct gtagcaccgc ctacatacct cgctctgcta atcctgttac cagtggctgc
     4261 tgccagtggc gataagtcgt gtcttaccgg gttggactca agacgatagt taccggataa
     4321 ggcgcagcgg tcgggctgaa cggggggttc gtgcacacag cccagcttgg agcgaacgac
     4381 ctacaccgaa ctgagatacc tacagcgtga gctatgagaa agcgccacgc ttcccgaagg
     4441 gagaaaggcg gacaggtatc cggtaagcgg cagggtcgga acaggagagc gcacgaggga
     4501 gcttccaggg ggaaacgcct ggtatcttta tagtcctgtc gggtttcgcc acctctgact
     4561 tgagcgtcga tttttgtgat gctcgtcagg ggggcggagc ctatggaaaa acgccagcaa
     4621 cgcggccttt ttacggttcc tggccttttg ctggcctttt gctcacatgt tctttcctgc
     4681 gttatcccct gattctgtgg ataaccgtat taccgccttt gagtgagctg ataccgctcg
     4741 ccgcagccga acgaccgagc gcagcgagtc agtgagcgag gaagcggaag agcgcccaat
     4801 acgcaaaccg cctctccccg cgcgttggcc gattcattaa tgcagctggc acgacaggtt
     4861 tcccgactgg aaagcgggca gtgagcgcaa cgcaattaat gtgagttagc tcactcatta
     4921 ggcaccccag gctttacact ttatgcttcc ggctcgtatg ttgtgtggaa ttgtgagcgg
     4981 ataacaattt cacacaggaa acagctatga ccatgattac gccaagctct agctagaggt
     5041 cgacggtata cagacatgat aagatacatt gatgagtttg gacaaaccac aactagaatg
     5101 cagtgaaaaa aatgctttat ttgtgaaatt tgtgatgcta ttgctttatt tgtaaccatt
     5161 ataagctgca ataaacaagt tggggtgggc gaagaactcc agcatgagat ccccgcgctg
     5221 gaggatcatc cagccggcgt cccggaaaac gattccgaag cccaaccttt catagaaggc
     5281 ggcggtggaa tcgaaatctc gtgatggcag gttgggcgtc gcttggtcgg tcatttcgcg
     5341 aaccccagag tcccgctcag aagaactcgt caagaaggcg atagaaggcg atgcgctgcg
     5401 aatcgggagc ggcgataccg taaagcacga ggaagcggtc agcccattcg ccgccaagct
     5461 cttcagcaat atcacgggta gccaacgcta tgtcctgata gcggtccgcc acacccagcc
     5521 ggccacagtc gatgaatcca gaaaagcggc cattttccac catgatattc ggcaagcagg
     5581 catcgccatg ggtcacgacg agatcctcgc cgtcgggcat gcgcgccttg agcctggcga
     5641 acagttcggc tggcgcgagc ccctgatgct cttcgtccag atcatcctga tcgacaagac
     5701 cggcttccat ccgagtacgt gctcgctcga tgcgatgttt cgcttggtgg tcgaatgggc
     5761 aggtagccgg atcaagcgta tgcagccgcc gcattgcatc agccatgatg gatactttct
     5821 cggcaggagc aaggtgagat gacaggagat cctgccccgg cacttcgccc aatagcagcc
     5881 agtcccttcc cgcttcagtg acaacgtcga gcacagctgc gcaaggaacg cccgtcgtgg
     5941 ccagccacga tagccgcgct gcctcgtcct gcagttcatt cagggcaccg gacaggtcgg
     6001 tcttgacaaa aagaaccggg cgcccctgcg ctgacagccg gaacacggcg gcatcagagc
     6061 agccgattgt ctgttgtgcc cagtcatagc cgaatagcct ctccacccaa gcggccggag
     6121 aacctgcgtg caatccatct tgttcaatca tgcgaaacga tcctcatcct gtctcttgat
     6181 cagatccgaa aatggatata caagctcccg ggagcttttt gcaaaagcct aggcctccaa
     6241 aaaagcctcc tcactacttc tggaatagct cagaggcaga ggcggcctcg gcctctgcat
     6301 aaataaaaaa aattagtcag ccatggggcg gagaatgggc ggaactgggc ggagttaggg
     6361 gcgggatggg cggagttagg ggcgggacta tggttgctga ctaattgaga tgcatgcttt
     6421 gcatacttct gcctgctggg gagcctgggg actttccaca cctggttgct gactaattga
     6481 gatgcatgct ttgcatactt ctgcctgctg gggagcctgg ggactttcca caccctaact
     6541 gacacacatt ccacagaatt aattcgcgtt aaatttttgt taaatcagct cattttttaa
     6601 ccaataggcc gaaatcggca aaatccctta taaatcaaaa gaatagaccg agatagggtt
     6661 gagtgttgtt ccagtttgga acaagagtcc actattaaag aacgtggact ccaacgtcaa
     6721 agggcgaaaa accgtctatc agggcgatgg cccactacgt gaaccatcac cctaatcaag
     6781 ttttttgggg tcgaggtgcc gtaaagcact aaatcggaac cctaaaggga gcccccgatt
     6841 tagagcttga cggggaaagc cggcgaacgt ggcgagaaag gaagggaaga aagcgaaagg
     6901 agcgggcgct agggcgctgg caagtgtagc ggtcacgctg cgcgtaacca ccacacccgc
     6961 cgcgcttaat gcgccgctac agggcgcgtg gggatacccc ctagagcccc agctggttct
     7021 ttccgcctca gaagccatag agcccaccgc atccccagca tgcctgctat tgtcttccca
     7081 atcctc
//
