LOCUS       sequence-199234-        7161 bp    DNA     circular SYN 15-DEC-2025
DEFINITION  Anion channelrhodopsin from Geminigera cryophila (CCMP2564) for 
            expression in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    .
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7161)
  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 7161)
  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-GcACR_145-EYFP
FEATURES             Location/Qualifiers
     source          1..7161
                     /mol_type="other DNA"
                     /organism="synthetic DNA construct"
     primer_bind     200..220
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     rep_origin      complement(229..657)
                     /direction=LEFT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(339..360)
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     551..570
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     polyA_signal    703..927
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     primer_bind     916..933
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     CDS             complement(956..973)
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     CDS             complement(983..1024)
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             complement(join(1054..1767,1768..1770))
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(1099..1120)
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     primer_bind     1448..1467
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     1707..1728
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
     primer_bind     complement(2801..2820)
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        complement(2802..2820)
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     promoter        complement(2865..3068)
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     complement(2894..2914)
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     enhancer        3069..3448
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     primer_bind     3620..3639
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     promoter        3714..3818
                     /gene="bla"
                     /label=AmpR promoter
     CDS             join(3819..3887,3888..4679)
                     /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(4037..4056)
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     rep_origin      4850..5438
                     /direction=RIGHT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     5339..5358
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     primer_bind     5592..5609
                     /label=L4440
                     /note="L4440 vector, forward primer"
     protein_bind    5726..5747
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     promoter        join(5762..5767,5768..5785,5786..5792)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    5800..5816
                     /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     5805..5827
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     primer_bind     5824..5840
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     5824..5840
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     polyA_signal    5889..6010
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(5900..5919)
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     5954..5973
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     CDS             complement(6186..6980)
                     /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(6297..6316)
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     primer_bind     6907..6926
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     promoter        complement(join(7047..7161,1..215))
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      join(7061..7161,1..35)
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     complement(7115..7134)
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
ORIGIN
        1 gaatgggcgg aactgggcgg agttaggggc gggatgggcg gagttagggg cgggactatg
       61 gttgctgact aattgagatg catgctttgc atacttctgc ctgctgggga gcctggggac
      121 tttccacacc tggttgctga ctaattgaga tgcatgcttt gcatacttct gcctgctggg
      181 gagcctgggg actttccaca ccctaactga cacacattcc acagaattaa ttcgcgttaa
      241 atttttgtta aatcagctca ttttttaacc aataggccga aatcggcaaa atcccttata
      301 aatcaaaaga atagaccgag atagggttga gtgttgttcc agtttggaac aagagtccac
      361 tattaaagaa cgtggactcc aacgtcaaag ggcgaaaaac cgtctatcag ggcgatggcc
      421 cactacgtga accatcaccc taatcaagtt ttttggggtc gaggtgccgt aaagcactaa
      481 atcggaaccc taaagggagc ccccgattta gagcttgacg gggaaagccg gcgaacgtgg
      541 cgagaaagga agggaagaaa gcgaaaggag cgggcgctag ggcgctggca agtgtagcgg
      601 tcacgctgcg cgtaaccacc acacccgccg cgcttaatgc gccgctacag ggcgcgtggg
      661 gataccccct agagccccag ctggttcttt ccgcctcaga agccatagag cccaccgcat
      721 ccccagcatg cctgctattg tcttcccaat cctccccctt gctgtcctgc cccaccccac
      781 cccccagaat agaatgacac ctactcagac aatgcgatgc aatttcctca ttttattagg
      841 aaaggacagt gggagtggca ccttccaggg tcaaggaagg cacgggggag gggcaaacaa
      901 cagatggctg gcaactagaa ggcacagtcg aggctgatca gcgggtttaa actcaatggt
      961 gatggtgatg atgaccggta cgcgtagaat cgagaccgag gagagggtta gggataggct
     1021 taccttcgaa ccgcgggccc tctagagcat atgttacttg tacagctcgt ccatgccgag
     1081 agtgatcccg gcggcggtca cgaactccag caggaccatg tgatcgcgct tctcgttggg
     1141 gtctttgctc agggcggact ggtagctcag gtagtggttg tcgggcagca gcacggggcc
     1201 gtcgccgatg ggggtgttct gctggtagtg gtcggcgagc tgcacgctgc cgtcctcgat
     1261 gttgtggcgg atcttgaagt tcaccttgat gccgttcttc tgcttgtcgg ccatgatata
     1321 gacgttgtgg ctgttgtagt tgtactccag cttgtgcccc aggatgttgc cgtcctcctt
     1381 gaagtcgatg cccttcagct cgatgcggtt caccagggtg tcgccctcga acttcacctc
     1441 ggcgcgggtc ttgtagttgc cgtcgtcctt gaagaagatg gtgcgctcct ggacgtagcc
     1501 ttcgggcatg gcggacttga agaagtcgtg ctgcttcatg tggtcggggt agcgggcgaa
     1561 gcactgcagg ccgtagccga aggtggtcac gagggtgggc cagggcacgg gcagcttgcc
     1621 ggtggtgcag atgaacttca gggtcagctt gccgtaggtg gcatcgccct cgccctcgcc
     1681 ggacacgctg aacttgtggc cgtttacgtc gccgtccagc tcgaccagga tgggcaccac
     1741 cccggtgaac agctcctcgc ccttgctcac ggcggccgcg gcgctgttga atctgccatg
     1801 aggggcggcg aaggtgtcct cctccttgtg gttgtggcta ttggcggggg cctcttcgtc
     1861 gtggttgttg gggtgcttct tgggggtcca tgttccgccc aggaggcccc acacggtgtt
     1921 ggtctggatg acgccataca ggttcttgca cagggcgtcg gcgcacaggt agcccagggc
     1981 ggatgtctgt tcggagatca cacagaggcc ggtggagctc agcacccaaa acacagggaa
     2041 agcggtccag gaggtgaaga agatgaacct catgaagtac agcctcctca gggctgtggc
     2101 gttttcaggg acgtcggtgg tgggggtgaa cttggcgatg ccctgctgga agatgaacca
     2161 cactgtggtg atctccatgg tgtagaagct gaagcccagg aagaagaaca gccacttggc
     2221 gatcttggtg tcggtggtgg tgctggccag caggccacac acggtcctga tcaggctggc
     2281 ggcgcagatg ggcatcctca gctggatgcc gtagagcttc atgtcgtgga tgccgaaagc
     2341 ctggctcagc atgatgggac aggtgatcag ccagctacac attctggaca ggggcagcat
     2401 tctgccgtcg cccatcctga ggactccgta tccggcgccg gtgcagcagt acaggatgca
     2461 ctccaccagg ggcaggtaca cggcttccca ggtcaccttg ggggccctgg tcatcatgaa
     2521 cagggcctga cagaggcaca ccacggccca gagtgtgatg attgttctct ccctgctggt
     2581 ggcgtgcagg cccttgttca cgcaatcggg gttggcgatc tcccactcgc cgtacatgcc
     2641 agcgtaaggg gtggcagggc cgatgccatc gatgtaggcg gcctttcccc tgaccaccat
     2701 tctctcttca ggctggaatc ccacggcttg gctgccggtg tcctgagcgt cgagcatgga
     2761 tccgagctcg gtaccaagct taactagcca gcttgggtct ccctatagtg agtcgtatta
     2821 atttcgataa gccagtaagc agtgggttct ctagttagcc agagagctct gcttatatag
     2881 acctcccacc gtacacgcct accgcccatt tgcgtcaatg gggcggagtt gttacgacat
     2941 tttggaaagt cccgttgatt ttggtgccaa aacaaactcc cattgacgtc aatggggtgg
     3001 agacttggaa atccccgtga gtcaaaccgc tatccacgcc cattgatgta ctgccaaaac
     3061 cgcatcacca tggtaatagc gatgactaat acgtagatgt actgccaagt aggaaagtcc
     3121 cataaggtca tgtactgggc ataatgccag gcgggccatt taccgtcatt gacgtcaata
     3181 gggggcgtac ttggcatatg atacacttga tgtactgcca agtgggcagt ttaccgtaaa
     3241 tactccaccc attgacgtca atggaaagtc cctattggcg ttactatggg aacatacgtc
     3301 attattgacg tcaatgggcg ggggtcgttg ggcggtcagc caggcgggcc atttaccgta
     3361 agttatgtaa cgcggaactc catatatggg ctatgaacta atgaccccgt aattgattac
     3421 tattaataac tagtcaataa tcaatgtcaa cgcgtatatc tggcccgtac atcgcgaagc
     3481 agcgcaaaac gcctaaccct aagcagattc ttcatgcaat tgtcggtcaa gccttgcctt
     3541 gttgtagctt aaattttgct cgcgcactac tcagcgacct ccaacacaca agcagggagc
     3601 agatactggc ttaactatgc ggcatcagag cagattgtac tgagagtgca ccatagggga
     3661 tcgggagatc tcccgatccg tcgacgtcag gtggcacttt tcggggaaat gtgcgcggaa
     3721 cccctatttg tttatttttc taaatacatt caaatatgta tccgctcatg agacaataac
     3781 cctgataaat gcttcaataa tattgaaaaa ggaagagtat gagtattcaa catttccgtg
     3841 tcgcccttat tccctttttt gcggcatttt gccttcctgt ttttgctcac ccagaaacgc
     3901 tggtgaaagt aaaagatgct gaagatcagt tgggtgcacg agtgggttac atcgaactgg
     3961 atctcaacag cggtaagatc cttgagagtt ttcgccccga agaacgtttt ccaatgatga
     4021 gcacttttaa agttctgcta tgtggcgcgg tattatcccg tattgacgcc gggcaagagc
     4081 aactcggtcg ccgcatacac tattctcaga atgacttggt tgagtactca ccagtcacag
     4141 aaaagcatct tacggatggc atgacagtaa gagaattatg cagtgctgcc ataaccatga
     4201 gtgataacac tgcggccaac ttacttctga caacgatcgg aggaccgaag gagctaaccg
     4261 cttttttgca caacatgggg gatcatgtaa ctcgccttga tcgttgggaa ccggagctga
     4321 atgaagccat accaaacgac gagcgtgaca ccacgatgcc tgtagcaatg gcaacaacgt
     4381 tgcgcaaact attaactggc gaactactta ctctagcttc ccggcaacaa ttaatagact
     4441 ggatggaggc ggataaagtt gcaggaccac ttctgcgctc ggcccttccg gctggctggt
     4501 ttattgctga taaatctgga gccggtgagc gtgggtctcg cggtatcatt gcagcactgg
     4561 ggccagatgg taagccctcc cgtatcgtag ttatctacac gacggggagt caggcaacta
     4621 tggatgaacg aaatagacag atcgctgaga taggtgcctc actgattaag cattggtaac
     4681 tgtcagacca agtttactca tatatacttt agattgattt aaaacttcat ttttaattta
     4741 aaaggatcta ggtgaagatc ctttttgata atctcatgac caaaatccct taacgtgagt
     4801 tttcgttcca ctgagcgtca gaccccgtag aaaagatcaa aggatcttct tgagatcctt
     4861 tttttctgcg cgtaatctgc tgcttgcaaa caaaaaaacc accgctacca gcggtggttt
     4921 gtttgccgga tcaagagcta ccaactcttt ttccgaaggt aactggcttc agcagagcgc
     4981 agataccaaa tactgttctt ctagtgtagc cgtagttagg ccaccacttc aagaactctg
     5041 tagcaccgcc tacatacctc gctctgctaa tcctgttacc agtggctgct gccagtggcg
     5101 ataagtcgtg tcttaccggg ttggactcaa gacgatagtt accggataag gcgcagcggt
     5161 cgggctgaac ggggggttcg tgcacacagc ccagcttgga gcgaacgacc tacaccgaac
     5221 tgagatacct acagcgtgag ctatgagaaa gcgccacgct tcccgaaggg agaaaggcgg
     5281 acaggtatcc ggtaagcggc agggtcggaa caggagagcg cacgagggag cttccagggg
     5341 gaaacgcctg gtatctttat agtcctgtcg ggtttcgcca cctctgactt gagcgtcgat
     5401 ttttgtgatg ctcgtcaggg gggcggagcc tatggaaaaa cgccagcaac gcggcctttt
     5461 tacggttcct ggccttttgc tggccttttg ctcacatgtt ctttcctgcg ttatcccctg
     5521 attctgtgga taaccgtatt accgcctttg agtgagctga taccgctcgc cgcagccgaa
     5581 cgaccgagcg cagcgagtca gtgagcgagg aagcggaaga gcgcccaata cgcaaaccgc
     5641 ctctccccgc gcgttggccg attcattaat gcagctggca cgacaggttt cccgactgga
     5701 aagcgggcag tgagcgcaac gcaattaatg tgagttagct cactcattag gcaccccagg
     5761 ctttacactt tatgcttccg gctcgtatgt tgtgtggaat tgtgagcgga taacaatttc
     5821 acacaggaaa cagctatgac catgattacg ccaagctcta gctagaggtc gacggtatac
     5881 agacatgata agatacattg atgagtttgg acaaaccaca actagaatgc agtgaaaaaa
     5941 atgctttatt tgtgaaattt gtgatgctat tgctttattt gtaaccatta taagctgcaa
     6001 taaacaagtt ggggtgggcg aagaactcca gcatgagatc cccgcgctgg aggatcatcc
     6061 agccggcgtc ccggaaaacg attccgaagc ccaacctttc atagaaggcg gcggtggaat
     6121 cgaaatctcg tgatggcagg ttgggcgtcg cttggtcggt catttcgcga accccagagt
     6181 cccgctcaga agaactcgtc aagaaggcga tagaaggcga tgcgctgcga atcgggagcg
     6241 gcgataccgt aaagcacgag gaagcggtca gcccattcgc cgccaagctc ttcagcaata
     6301 tcacgggtag ccaacgctat gtcctgatag cggtccgcca cacccagccg gccacagtcg
     6361 atgaatccag aaaagcggcc attttccacc atgatattcg gcaagcaggc atcgccatgg
     6421 gtcacgacga gatcctcgcc gtcgggcatg cgcgccttga gcctggcgaa cagttcggct
     6481 ggcgcgagcc cctgatgctc ttcgtccaga tcatcctgat cgacaagacc ggcttccatc
     6541 cgagtacgtg ctcgctcgat gcgatgtttc gcttggtggt cgaatgggca ggtagccgga
     6601 tcaagcgtat gcagccgccg cattgcatca gccatgatgg atactttctc ggcaggagca
     6661 aggtgagatg acaggagatc ctgccccggc acttcgccca atagcagcca gtcccttccc
     6721 gcttcagtga caacgtcgag cacagctgcg caaggaacgc ccgtcgtggc cagccacgat
     6781 agccgcgctg cctcgtcctg cagttcattc agggcaccgg acaggtcggt cttgacaaaa
     6841 agaaccgggc gcccctgcgc tgacagccgg aacacggcgg catcagagca gccgattgtc
     6901 tgttgtgccc agtcatagcc gaatagcctc tccacccaag cggccggaga acctgcgtgc
     6961 aatccatctt gttcaatcat gcgaaacgat cctcatcctg tctcttgatc agatccgaaa
     7021 atggatatac aagctcccgg gagctttttg caaaagccta ggcctccaaa aaagcctcct
     7081 cactacttct ggaatagctc agaggcagag gcggcctcgg cctctgcata aataaaaaaa
     7141 attagtcagc catggggcgg a
//
