LOCUS       Exported                7092 bp ds-DNA     circular SYN 13-MAY-2021
DEFINITION  Anion channelrhodopsin from Rhodomonas sp. (CCMP768) for expression 
            in mammalian cells.
ACCESSION   .
VERSION     .
KEYWORDS    pcDNA3.1-R1ACR_799-EYFP
SOURCE      synthetic DNA construct
  ORGANISM  synthetic DNA construct
REFERENCE   1  (bases 1 to 7092)
  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 7092)
  AUTHORS   .
  TITLE     Direct Submission
  JOURNAL   Exported May 13, 2021 from SnapGene Server 1.1.58
            http://www.snapgene.com
FEATURES             Location/Qualifiers
     source          1..7092
                     /organism="synthetic DNA construct"
                     /mol_type="other DNA"
     primer_bind     complement(156..175)
                     /label=EXFP-R
                     /note="For distinguishing EGFP variants, reverse primer"
     primer_bind     503..524
                     /label=EGFP-C
                     /note="EGFP, forward primer"
     CDS             599..640
                     /codon_start=1
                     /product="epitope tag from simian virus 5"
                     /label=V5 tag
                     /translation="GKPIPNPLLGLDST"
     CDS             650..667
                     /codon_start=1
                     /product="6xHis affinity tag"
                     /label=6xHis
                     /translation="HHHHHH"
     primer_bind     complement(690..707)
                     /label=BGH-rev
                     /note="Bovine growth hormone terminator, reverse primer. 
                     Also called BGH reverse"
     polyA_signal    696..920
                     /label=bGH poly(A) signal
                     /note="bovine growth hormone polyadenylation signal"
     rep_origin      966..1394
                     /direction=RIGHT
                     /label=f1 ori
                     /note="f1 bacteriophage origin of replication; arrow 
                     indicates direction of (+) strand synthesis"
     primer_bind     complement(1053..1072)
                     /label=F1ori-R
                     /note="F1 origin, reverse primer"
     primer_bind     1263..1284
                     /label=F1ori-F
                     /note="F1 origin, forward primer"
     primer_bind     complement(1403..1423)
                     /label=pBABE 3'
                     /note="SV40 enhancer, reverse primer for pBABE vectors"
     promoter        1408..1737
                     /label=SV40 promoter
                     /note="SV40 enhancer and early promoter"
     rep_origin      1588..1723
                     /label=SV40 ori
                     /note="SV40 origin of replication"
     primer_bind     1650..1669
                     /label=SV40pro-F
                     /note="SV40 promoter/origin, forward primer"
     CDS             1804..2598
                     /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(1858..1877)
                     /label=Neo-R
                     /note="Neomycin resistance gene, reverse primer"
     primer_bind     2468..2487
                     /label=Neo-F
                     /note="Neomycin resistance gene, forward primer"
     polyA_signal    2774..2895
                     /label=SV40 poly(A) signal
                     /note="SV40 polyadenylation signal"
     primer_bind     complement(2811..2830)
                     /label=SV40pA-R
                     /note="SV40 polyA, reverse primer"
     primer_bind     2865..2884
                     /label=EBV-rev
                     /note="SV40 polyA terminator, reverse primer"
     primer_bind     complement(2944..2960)
                     /label=M13 rev
                     /note="common sequencing primer, one of multiple similar 
                     variants"
     primer_bind     complement(2944..2960)
                     /label=M13 Reverse
                     /note="In lacZ gene. Also called M13-rev"
     primer_bind     complement(2957..2979)
                     /label=M13/pUC Reverse
                     /note="In lacZ gene"
     protein_bind    2968..2984
                     /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(2992..3022)
                     /label=lac promoter
                     /note="promoter for the E. coli lac operon"
     protein_bind    3037..3058
                     /label=CAP binding site
                     /bound_moiety="E. coli catabolite activator protein"
                     /note="CAP binding activates transcription in the presence 
                     of cAMP."
     primer_bind     complement(3175..3192)
                     /label=L4440
                     /note="L4440 vector, forward primer"
     rep_origin      complement(3346..3934)
                     /direction=LEFT
                     /label=ori
                     /note="high-copy-number ColE1/pMB1/pBR322/pUC origin of 
                     replication"
     primer_bind     complement(3426..3445)
                     /label=pBR322ori-F
                     /note="pBR322 origin, forward primer"
     CDS             complement(4105..4965)
                     /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     4728..4747
                     /label=Amp-R
                     /note="Ampicillin resistance gene, reverse primer"
     promoter        complement(4966..5070)
                     /gene="bla"
                     /label=AmpR promoter
     primer_bind     complement(5145..5164)
                     /label=pRS-marker
                     /note="pRS vectors, use to sequence yeast selectable 
                     marker"
     enhancer        5336..5715
                     /label=CMV enhancer
                     /note="human cytomegalovirus immediate early enhancer"
     promoter        5716..5919
                     /label=CMV promoter
                     /note="human cytomegalovirus (CMV) immediate early 
                     promoter"
     primer_bind     5870..5890
                     /label=CMV-F
                     /note="Human CMV immediate early promoter, forward primer"
     primer_bind     5964..5983
                     /label=T7
                     /note="T7 promoter, forward primer"
     promoter        5964..5982
                     /label=T7 promoter
                     /note="promoter for bacteriophage T7 RNA polymerase"
     CDS             6945..569
                     /codon_start=1
                     /product="enhanced YFP"
                     /label=EYFP
                     /note="mammalian codon-optimized"
                     /translation="VSKGEELFTGVVPILVELDGDVNGHKFSVSGEGEGDATYGKLTLK
                     FICTTGKLPVPWPTLVTTFGYGLQCFARYPDHMKQHDFFKSAMPEGYVQERTIFFKDDG
                     NYKTRAEVKFEGDTLVNRIELKGIDFKEDGNILGHKLEYNYNSHNVYIMADKQKNGIKV
                     NFKIRHNIEDGSVQLADHYQQNTPIGDGPVLLPDNHYLSYQSALSKDPNEKRDHMVLLE
                     FVTAAGITLGMDELYK"
     primer_bind     complement(6987..7008)
                     /label=EGFP-N
                     /note="EGFP, reverse primer"
ORIGIN
        1 ccggcaagct gcccgtgccc tggcccaccc tcgtgaccac cttcggctac ggcctgcagt
       61 gcttcgcccg ctaccccgac cacatgaagc agcacgactt cttcaagtcc gccatgcccg
      121 aaggctacgt ccaggagcgc accatcttct tcaaggacga cggcaactac aagacccgcg
      181 ccgaggtgaa gttcgagggc gacaccctgg tgaaccgcat cgagctgaag ggcatcgact
      241 tcaaggagga cggcaacatc ctggggcaca agctggagta caactacaac agccacaacg
      301 tctatatcat ggccgacaag cagaagaacg gcatcaaggt gaacttcaag atccgccaca
      361 acatcgagga cggcagcgtg cagctcgccg accactacca gcagaacacc cccatcggcg
      421 acggccccgt gctgctgccc gacaaccact acctgagcta ccagtccgcc ctgagcaaag
      481 accccaacga gaagcgcgat cacatggtcc tgctggagtt cgtgaccgcc gccgggatca
      541 ctctcggcat ggacgagctg tacaagtaac atatgctcta gagggcccgc ggttcgaagg
      601 taagcctatc cctaaccctc tcctcggtct cgattctacg cgtaccggtc atcatcacca
      661 tcaccattga gtttaaaccc gctgatcagc ctcgactgtg ccttctagtt gccagccatc
      721 tgttgtttgc ccctcccccg tgccttcctt gaccctggaa ggtgccactc ccactgtcct
      781 ttcctaataa aatgaggaaa ttgcatcgca ttgtctgagt aggtgtcatt ctattctggg
      841 gggtggggtg gggcaggaca gcaaggggga ggattgggaa gacaatagca ggcatgctgg
      901 ggatgcggtg ggctctatgg cttctgaggc ggaaagaacc agctggggct ctagggggta
      961 tccccacgcg ccctgtagcg gcgcattaag cgcggcgggt gtggtggtta cgcgcagcgt
     1021 gaccgctaca cttgccagcg ccctagcgcc cgctcctttc gctttcttcc cttcctttct
     1081 cgccacgttc gccggctttc cccgtcaagc tctaaatcgg gggctccctt tagggttccg
     1141 atttagtgct ttacggcacc tcgaccccaa aaaacttgat tagggtgatg gttcacgtag
     1201 tgggccatcg ccctgataga cggtttttcg ccctttgacg ttggagtcca cgttctttaa
     1261 tagtggactc ttgttccaaa ctggaacaac actcaaccct atctcggtct attcttttga
     1321 tttataaggg attttgccga tttcggccta ttggttaaaa aatgagctga tttaacaaaa
     1381 atttaacgcg aattaattct gtggaatgtg tgtcagttag ggtgtggaaa gtccccaggc
     1441 tccccagcag gcagaagtat gcaaagcatg catctcaatt agtcagcaac caggtgtgga
     1501 aagtccccag gctccccagc aggcagaagt atgcaaagca tgcatctcaa ttagtcagca
     1561 accatagtcc cgcccctaac tccgcccatc ccgcccctaa ctccgcccag ttccgcccat
     1621 tctccgcccc atggctgact aatttttttt atttatgcag aggccgaggc cgcctctgcc
     1681 tctgagctat tccagaagta gtgaggaggc ttttttggag gcctaggctt ttgcaaaaag
     1741 ctcccgggag cttgtatatc cattttcgga tctgatcaag agacaggatg aggatcgttt
     1801 cgcatgattg aacaagatgg attgcacgca ggttctccgg ccgcttgggt ggagaggcta
     1861 ttcggctatg actgggcaca acagacaatc ggctgctctg atgccgccgt gttccggctg
     1921 tcagcgcagg ggcgcccggt tctttttgtc aagaccgacc tgtccggtgc cctgaatgaa
     1981 ctgcaggacg aggcagcgcg gctatcgtgg ctggccacga cgggcgttcc ttgcgcagct
     2041 gtgctcgacg ttgtcactga agcgggaagg gactggctgc tattgggcga agtgccgggg
     2101 caggatctcc tgtcatctca ccttgctcct gccgagaaag tatccatcat ggctgatgca
     2161 atgcggcggc tgcatacgct tgatccggct acctgcccat tcgaccacca agcgaaacat
     2221 cgcatcgagc gagcacgtac tcggatggaa gccggtcttg tcgatcagga tgatctggac
     2281 gaagagcatc aggggctcgc gccagccgaa ctgttcgcca ggctcaaggc gcgcatgccc
     2341 gacggcgagg atctcgtcgt gacccatggc gatgcctgct tgccgaatat catggtggaa
     2401 aatggccgct tttctggatt catcgactgt ggccggctgg gtgtggcgga ccgctatcag
     2461 gacatagcgt tggctacccg tgatattgct gaagagcttg gcggcgaatg ggctgaccgc
     2521 ttcctcgtgc tttacggtat cgccgctccc gattcgcagc gcatcgcctt ctatcgcctt
     2581 cttgacgagt tcttctgagc gggactctgg ggttcgcgaa atgaccgacc aagcgacgcc
     2641 caacctgcca tcacgagatt tcgattccac cgccgccttc tatgaaaggt tgggcttcgg
     2701 aatcgttttc cgggacgccg gctggatgat cctccagcgc ggggatctca tgctggagtt
     2761 cttcgcccac cccaacttgt ttattgcagc ttataatggt tacaaataaa gcaatagcat
     2821 cacaaatttc acaaataaag catttttttc actgcattct agttgtggtt tgtccaaact
     2881 catcaatgta tcttatcatg tctgtatacc gtcgacctct agctagagct tggcgtaatc
     2941 atggtcatag ctgtttcctg tgtgaaattg ttatccgctc acaattccac acaacatacg
     3001 agccggaagc ataaagtgta aagcctgggg tgcctaatga gtgagctaac tcacattaat
     3061 tgcgttgcgc tcactgcccg ctttccagtc gggaaacctg tcgtgccagc tgcattaatg
     3121 aatcggccaa cgcgcgggga gaggcggttt gcgtattggg cgctcttccg cttcctcgct
     3181 cactgactcg ctgcgctcgg tcgttcggct gcggcgagcg gtatcagctc actcaaaggc
     3241 ggtaatacgg ttatccacag aatcagggga taacgcagga aagaacatgt gagcaaaagg
     3301 ccagcaaaag gccaggaacc gtaaaaaggc cgcgttgctg gcgtttttcc ataggctccg
     3361 cccccctgac gagcatcaca aaaatcgacg ctcaagtcag aggtggcgaa acccgacagg
     3421 actataaaga taccaggcgt ttccccctgg aagctccctc gtgcgctctc ctgttccgac
     3481 cctgccgctt accggatacc tgtccgcctt tctcccttcg ggaagcgtgg cgctttctca
     3541 tagctcacgc tgtaggtatc tcagttcggt gtaggtcgtt cgctccaagc tgggctgtgt
     3601 gcacgaaccc cccgttcagc ccgaccgctg cgccttatcc ggtaactatc gtcttgagtc
     3661 caacccggta agacacgact tatcgccact ggcagcagcc actggtaaca ggattagcag
     3721 agcgaggtat gtaggcggtg ctacagagtt cttgaagtgg tggcctaact acggctacac
     3781 tagaagaaca gtatttggta tctgcgctct gctgaagcca gttaccttcg gaaaaagagt
     3841 tggtagctct tgatccggca aacaaaccac cgctggtagc ggtggttttt ttgtttgcaa
     3901 gcagcagatt acgcgcagaa aaaaaggatc tcaagaagat cctttgatct tttctacggg
     3961 gtctgacgct cagtggaacg aaaactcacg ttaagggatt ttggtcatga gattatcaaa
     4021 aaggatcttc acctagatcc ttttaaatta aaaatgaagt tttaaatcaa tctaaagtat
     4081 atatgagtaa acttggtctg acagttacca atgcttaatc agtgaggcac ctatctcagc
     4141 gatctgtcta tttcgttcat ccatagttgc ctgactcccc gtcgtgtaga taactacgat
     4201 acgggagggc ttaccatctg gccccagtgc tgcaatgata ccgcgagacc cacgctcacc
     4261 ggctccagat ttatcagcaa taaaccagcc agccggaagg gccgagcgca gaagtggtcc
     4321 tgcaacttta tccgcctcca tccagtctat taattgttgc cgggaagcta gagtaagtag
     4381 ttcgccagtt aatagtttgc gcaacgttgt tgccattgct acaggcatcg tggtgtcacg
     4441 ctcgtcgttt ggtatggctt cattcagctc cggttcccaa cgatcaaggc gagttacatg
     4501 atcccccatg ttgtgcaaaa aagcggttag ctccttcggt cctccgatcg ttgtcagaag
     4561 taagttggcc gcagtgttat cactcatggt tatggcagca ctgcataatt ctcttactgt
     4621 catgccatcc gtaagatgct tttctgtgac tggtgagtac tcaaccaagt cattctgaga
     4681 atagtgtatg cggcgaccga gttgctcttg cccggcgtca atacgggata ataccgcgcc
     4741 acatagcaga actttaaaag tgctcatcat tggaaaacgt tcttcggggc gaaaactctc
     4801 aaggatctta ccgctgttga gatccagttc gatgtaaccc actcgtgcac ccaactgatc
     4861 ttcagcatct tttactttca ccagcgtttc tgggtgagca aaaacaggaa ggcaaaatgc
     4921 cgcaaaaaag ggaataaggg cgacacggaa atgttgaata ctcatactct tcctttttca
     4981 atattattga agcatttatc agggttattg tctcatgagc ggatacatat ttgaatgtat
     5041 ttagaaaaat aaacaaatag gggttccgcg cacatttccc cgaaaagtgc cacctgacgt
     5101 cgacggatcg ggagatctcc cgatccccta tggtgcactc tcagtacaat ctgctctgat
     5161 gccgcatagt taagccagta tctgctccct gcttgtgtgt tggaggtcgc tgagtagtgc
     5221 gcgagcaaaa tttaagctac aacaaggcaa ggcttgaccg acaattgcat gaagaatctg
     5281 cttagggtta ggcgttttgc gctgcttcgc gatgtacggg ccagatatac gcgttgacat
     5341 tgattattga ctagttatta atagtaatca attacggggt cattagttca tagcccatat
     5401 atggagttcc gcgttacata acttacggta aatggcccgc ctggctgacc gcccaacgac
     5461 ccccgcccat tgacgtcaat aatgacgtat gttcccatag taacgccaat agggactttc
     5521 cattgacgtc aatgggtgga gtatttacgg taaactgccc acttggcagt acatcaagtg
     5581 tatcatatgc caagtacgcc ccctattgac gtcaatgacg gtaaatggcc cgcctggcat
     5641 tatgcccagt acatgacctt atgggacttt cctacttggc agtacatcta cgtattagtc
     5701 atcgctatta ccatggtgat gcggttttgg cagtacatca atgggcgtgg atagcggttt
     5761 gactcacggg gatttccaag tctccacccc attgacgtca atgggagttt gttttggcac
     5821 caaaatcaac gggactttcc aaaatgtcgt aacaactccg ccccattgac gcaaatgggc
     5881 ggtaggcgtg tacggtggga ggtctatata agcagagctc tctggctaac tagagaaccc
     5941 actgcttact ggcttatcga aattaatacg actcactata gggagaccca agctggctag
     6001 ttaagcttgg taccgagctc ggatccatgg ccggcgtgca ggatgactgg agcgctgtga
     6061 gcgtgcctgc cttcggccag tatggcacat gggaagctgc caaccctgag tgcgtcgaga
     6121 tgggcctgag caccagcaag aacaccaaga tgatccagct gctgttcgcc agcatttgtg
     6181 cctgtcaggt gctgttcatg gcctccagag cccccagagt gacctgggag gccgtgtacc
     6241 tgcccctggt ggaaaccatc ctgtatggca tggcctccac cggcaacggc tccatgaggc
     6301 tggctgacgg cagaatggtg cccatcgcca ggttctgcag ctggatcatc acctgcccca
     6361 tcatgctctt tcaggtggtg ggcattcacg aggtgaaagt gtacggcatt tcctgcaaaa
     6421 acatggtgat ggccgccagc ctggtgagga ccgtgtacgg catcggagcc tccgtcacca
     6481 tcgacgacaa aatgaggtgg attcaataca tgctgtcctg cttcaccttc ctgttcgagc
     6541 tgtggtgcgt gtaccagatc tacaacgccg gcctgaagaa gttcagcgcc gtcaagaccc
     6601 ccctgaataa catcgtgcac tcccgtattc aattcctgag gttcctcttc tacctgagct
     6661 ggagcagctt ccccatcgtg tgggtgctga gcagcaccgg actgtgcctg atcaacgagg
     6721 acggctccgt gctggcttac ctggtcgccg atatcctgtg caagaacacc tacggaatca
     6781 tcaacgctaa taccctgttc agggtgctgg atggcaaatg gacacccgag accctggagg
     6841 aggccgccaa cacctacggc attatcaacg ccaacaccct gtttagagtg ctgaacggaa
     6901 agtgggagcc cgagaccgag gaggaagcta aaaacgcggc cgccgtgagc aagggcgagg
     6961 agctgttcac cggggtggtg cccatcctgg tcgagctgga cggcgacgta aacggccaca
     7021 agttcagcgt gtccggcgag ggcgagggcg atgccaccta cggcaagctg accctgaagt
     7081 tcatctgcac ca
//
